Check out our New Publishers' Select for Free Articles

Amino Acids: The Forum for Amino Acid, Peptide and Protein Research (v.42, #6)

Serotonin and molecular neuroimaging in humans using PET by Anne Saulin; Markus Savli; Rupert Lanzenberger (pp. 2039-2057).

The serotonergic system is one of the most important modulatory neurotransmitter systems in the human brain. It plays a central role in major physiological processes and is implicated in a number of psychiatric disorders. Along with the dopaminergic system, it is also one of the phylogenetically oldest human neurotransmitter systems and one of the most diverse, with 14 different receptors identified up to this day, many of whose function remains to be understood. The system’s functioning is even more diverse than the number of its receptors, since each is implicated in a number of different processes. This review aims at illustrating the distribution and summarizing the main functions of the serotonin (5-hydroxytryptamin, 5-HT) receptors as well as the serotonin transporter (SERT, 5-HTT), the vesicular monoamine transporter 2, monoamine oxidase type A and 5-HT synthesis in the human brain. Recent advances in in vivo quantification of these different receptors and enzymes that are part of the serotonergic system using positron emission tomography are described.

Keywords: Serotonin; PET; In vivo; Radioligand; Human brain; Neuroimaging; Molecular imaging

Changes in transcription during recovery from heat injury in Salmonella typhimurium and effects of BCAA on recovery by Wen Hsu-Ming; Kimitaka Naito; Yoshimasa Kinoshita; Hiroshi Kobayashi; Ken-ichi Honjoh; Kousuke Tashiro; Takahisa Miyamoto (pp. 2059-2066).

Mechanisms of recovery from heat injury in Salmonella typhimurium were elucidated. Recovery of the heat-injured S. typhimurium cells in TSB resulted in full recovery after 3 h of incubation at 37°C. The DNA microarray analysis of 30- and 60-min recovering cells resulted in an increase in transcription of 89 and 141 genes, respectively. Among them, 15 genes, with known function, seemed to be somewhat involved in recovery. They encoded proteins involved in branched-chain amino acid (BCAA) transport (livJ, livH), cell envelope integrity (ddg), heat-shock response (cpxP, rrmJ), phage shock protein (pspA), ribosome modulation factor (rmf), virulence (sseB) transcriptional regulation (rpoE, rpoH, rseA, rseB, rseC) and ArcB signal transduction (sixA) and cytoplasmic membrane protein (fxsA). Among them, the effects of BCAA supplementation on recovery from heat injury were studied to confirm the importance of the BCAA transport liv genes during recovery. It was found that supplementation of TSB with 0.1% BCAA resulted in an enhanced recovery of injured cells in comparison to those recovered in TSB without BCAA. Supplementation of BCAA at 0.1% resulted in a cell count increase 4.4-fold greater than that of the control after 1 h incubation. It seems that BCAA promoted the recovery by promoting protein synthesis either directly through their use in translation or indirectly through stimulation of protein synthesis by activation of the Lrp protein.

Keywords: Salmonella ; Heat injury; Recovery; BCAA; DNA microarray

Peptides derived from Mycobacterium tuberculosis Rv2301 protein are involved in invasion to human epithelial cells and macrophages by M. Ocampo; D. M. Rodríguez; H. Curtidor; M. Vanegas; M. A. Patarroyo; M. E. Patarroyo (pp. 2067-2077).

The specific function of putative cut2 protein (or CFP25), encoded by the Rv2301 gene from Mycobacterium tuberculosis H37Rv, has not been identified yet. The aim of this study was to assess some of CFP25 characteristics and its possible biological role in Mycobacterium tuberculosis H37Rv invasion process to target cells. Molecular assays indicated that the gene encoding Rv2301 is present and transcribed in M. tuberculosis complex strains. The presence of Rv2301 protein over the bacilli surface was confirmed by Western blot and immunoelectron microscopy analyses, using goats sera inoculated with synthetic peptides derived from Rv2301 protein. Receptor–ligand binding assays with carcinomic human alveolar basal epithelial cells (A549) and macrophages derived from human histolytic lymphoma monocytes (U937) allowed us to identify five high activity binding peptides (HABPs) in both cell lines, and two additional HABPs only in A549 cells. U937 HABPs binding interactions were characterized by saturation assays, finding dissociation constants (K _d) within the nanomolar range and positive cooperativity (n _H > 1). Inhibition assays were performed to assess the possible biological role of Rv2301 identified HABPs, finding that some of them were able to inhibit invasion at a 5 μM concentration, compared with the cytochalasin control. On the other hand, HABPs, and especially HABP 36507 located at the N-terminus of the protein, facilitated the internalization of fluorescent latex beads into A549 cells. These findings are of vital importance for the rational selection of Rv2301 HABPs, to be included as components of an antituberculosis vaccine.

Keywords: Mycobacterium tuberculosis ; High activity binding peptides; Cutinase; Rv2301

Influence of TiO₂ on prebiotic thermal synthesis of the Gly-Gln polymer by P. Leyton; R. Saladino; C. Crestini; M. Campos-Vallette; C. Paipa; A. Berríos; S. Fuentes; R. A. Zárate (pp. 2079-2088).

The role of the titanium dioxide (rutile and anatase) with and without room light on the thermal synthesis of the glycine–l-glutamine (Gly-Gln) polymer is described. The efficiency in percentage of polymerization with room light was increased in 6% in the presence of rutile and in 23% in the presence of anatase. The thermal synthesis in the molten state was carried out in the absence and presence of both oxides. In all cases, the vibrational spectra showed characteristic group frequencies corresponding to a polypeptide structure. No spectral differences were observed by room light effect on the polymer on rutile. However, the polymer obtained in the presence of anatase and room light shows spectral changes associated with the formation of shorter new abundant and conformationally different species compared with the original polymer. The SEM-EDX characterization of the solid phase involved in the thermal synthesis showed that the morphology of the polypeptide is different in the presence of rutile compared to anatase. The SDS–PAGE and GPC results suggest that smaller chains are formed in the presence of both oxides and the distribution of the size and weight of each polymer molecule is completely different when the condensation is performed in the presence of anatase or rutile. Nuclear magnetic resonance analyses confirmed the incorporation of both Gly and Gln residues in the polymers, with a prevalence of Gly. Both possible sequences N-GlyGln-C and N-GlnGly-C were also detected.

Keywords: Prebiotic chemistry; Thermal synthesis; Vibrational spectroscopy; Titanium dioxide

Effect of dietary arginine supplementation on reproductive performance of mice with porcine circovirus type 2 infection by Wenkai Ren; Yulong Yin; Gang Liu; Xinglong Yu; Yinghui Li; Guan Yang; Teijun Li; Guoyao Wu (pp. 2089-2094).

The objective of this study was to investigate whether supplemental dietary arginine increases reproductive performance in mice infected with porcine circovirus type2 (PCV2). A total of 50KM female mice were allotted randomly to the arginine group (0.6% arginine + gestation diet) and control group (1.22% alanine + gestation diet). All the mice began to mate after 14 days of treatment with our prepared feed and challenged with PCV2 at the dose of 100 TCID₅₀ (50% tissue culture infection dose, TCID₅₀) after 7 days of pregnancy. Abortion rate, litter number, litter birth weight, the daily weight gain in the first 7 days and survival rate in the first 2 weeks of the neonates were calculated. The serum progesterone, estrogen, nitric oxide and superoxide dismutase (SOD) activity and total antioxidant capacity (T-AOC) on the 14th day of pregnancy were measured. Arginine supplementation decreased the abortion rate of pregnant mice and mortality of neonates caused by PCV2 infection. Further, litter number, litter birth weight and the daily weight gain of neonates increased in the arginine group compared to the control group. Arginine supplementation increased significantly the serum progesterone (P < 0.01) and nitric oxide levels (P < 0.05), but had little effect on the serum estrogen level. SOD activity and T-AOC in the arginine group were significantly higher (P < 0.01) than the control group. In conclusion, arginine supplementation partially reversed the reproductive failure in mice caused by PCV2 infection.

Keywords: Arginine; Porcine circovirus; Progesterone; Nitric oxide

Metabolomic analysis of sulfur-containing substances and polyamines in regenerating rat liver by Young S. Jung; Sun J. Kim; Do Y. Kwon; Young C. Kim (pp. 2095-2102).

We studied the significance of alterations in the metabolomics of sulfur-containing substances in rapidly regenerating rat livers. Male rats were subjected to two-thirds partial hepatectomy (PHx), and the changes in hepatic levels of major sulfur-containing amino acids and related substances were monitored for 2 weeks. Liver weight began to increase from 24 h after the surgery, and appeared to recover fully in 2 weeks. Serum alanine aminotransferase and aspartate aminotransferase activities were elevated immediately after the surgery and returned slowly to normal levels in 2 weeks. Methionine, S-adenosylmethionine (SAM), cystathionine and cysteine were increased rapidly and remained elevated for longer than 1 week. Hepatic glutathione concentration was increased gradually for 24 h, and then decreased thereafter, whereas hypotaurine was elevated drastically right after the surgery. Hepatic concentrations of polyamines were altered significantly by PHx. In the hepatectomized livers putrescine concentration was elevated rapidly, reaching a level 40- to 50-fold greater than normal in 6–12 h. Ornithine, the metabolic substrate for putrescine synthesis, was also elevated markedly. Spermidine was increased significantly, whereas spermine was depressed below normal, which appeared to be due to the increased consumption of decarboxylated SAM for spermidine biosynthesis. The results show that the metabolomics of sulfur-containing amino acids and related substances is altered profoundly in regenerating rat livers until the original weight is recovered. Hepatic concentrations of polyamines after PHx are closely associated with the alteration in the metabolomics of sulfur-containing substances. The implication of these changes in the progression of liver regeneration is discussed.

Keywords: Sulfur-containing amino acids; Hepatectomy; Liver regeneration; Polyamines; S-adenosylmethionine; Glutathione

Evaluation of functional groups on amino acids in cyclic tetrapeptides in histone deacetylase inhibition by Md. Shahidul Islam; Mohammed P. I. Bhuiyan; Md. Nurul Islam; Tienabe Kipassa Nsiama; Naoto Oishi; Tamaki Kato; Norikazu Nishino; Akihiro Ito; Minoru Yoshida (pp. 2103-2110).

The naturally occurring cyclic tetrapeptide, chlamydocin, originally isolated from fungus Diheterospora chlamydosphoria, consists of α-aminoisobutyric acid, l-phenylalanine, d-proline and an unusual amino acid (S)-2-amino-8-((S)-oxiran-2-yl)-8-oxooctanoic acid (Aoe) and inhibits the histone deacetylases (HDACs), a class of regulatory enzymes. The epoxyketone moiety of Aoe is the key functional group for inhibition. The cyclic tetrapeptide scaffold is supposed to play important role for effective binding to the surface of enzymes. In place of the epoxyketone group, hydroxamic acid and sulfhydryl group have been applied to design inhibitor ligands to zinc atom in catalytic site of HDACs. In the research for more potent HDAC inhibitors, we replaced the epoxyketone moiety of Aoe with different functional groups and synthesized a series of chlamydocin analogs as HDAC inhibitors. Among the functional groups, methoxymethylketone moiety showed as potent inhibition as the hydroxamic acid. On the contrary, we confirmed that borate, trifruoromethylketone, and 2-aminoanilide are almost inactive in HDAC inhibition.

Keywords: Histone deacetylase inhibitor; Chlamydocin; Epoxyketone; Functional group design

Effects of dietary l-arginine or N-carbamylglutamate supplementation during late gestation of sows on the miR-15b/16, miR-221/222, VEGFA and eNOS expression in umbilical vein by X. D. Liu; X. Wu; Y. L. Yin; Y. Q. Liu; M. M. Geng; H. S. Yang; Francois Blachier; G. Y. Wu (pp. 2111-2119).

Placental vascular formation and blood flow are crucial for fetal survival, growth and development, and arginine regulates vascular development and function. This study determined the effects of dietary arginine or N-carbamylglutamate (NCG) supplementation during late gestation of sows on the microRNAs, vascular endothelial growth factor A (VEGFA) and endothelial nitric oxide synthase (eNOS) expression in umbilical vein. Twenty-seven landrace × large white sows at day (d) 90 of gestation were assigned randomly to three groups and fed the following diets: a control diet and the control diet supplemented with 1.0% l-arginine or 0.10% NCG. Umbilical vein of fetuses with body weight around 2.0 kg (oversized), 1.5 kg (normal) and 0.6 kg (intrauterine growth restriction, IUGR) were obtained immediately after farrowing for miR-15b, miR-16, miR-221, miR-222, VEGFA and eNOS real-time PCR analysis. Compared with the control diets, dietary Arg or NCG supplementation enhanced the reproductive performance of sows, significantly increased (P < 0.05) plasma arginine and decreased plasma VEGF and eNOS (P < 0.05). The miR-15b expression in the umbilical vein was higher (P < 0.05) in the NCG-supplemented group than in the control group. There was a trend in that the miR-222 expression in the umbilical vein of the oversized fetuses was higher (0.05 < P < 0.1) than in the normal and IUGR fetuses. The expression of eNOS in both Arg-supplemented and NCG-supplemented group were lower (P < 0.05) than in the control group. The expression of VEGFA was higher (P < 0.05) in the NCG-supplemented group than in the Arg-supplemented and the control group. Meanwhile, the expression of VEGFA of the oversized fetuses was higher (P < 0.05) than the normal and IUGR fetuses. In conclusion, this study demonstrated that dietary Arg or NCG supplementation may affect microRNAs (miR-15b, miR-222) targeting VEGFA and eNOS gene expressions in umbilical vein, so as to regulate the function and volume of the umbilical vein, provide more nutrients and oxygen from the maternal to the fetus tissue for fetal development and survival, and enhance the reproductive performance of sows.

Keywords: l-Arginine; N-Carbamylglutamate; Umbilical vein; VEGFA; ENOS; MicroRNAs

Efficient synthesis of unnatural dipeptides based on cis-2,5-disubstituted pyrrolidine by Ping-An Wang; Wei He; Si-Kun Cheng; Sheng-Yong Zhang (pp. 2121-2127).

The well-defined unnatural dipeptides based on cis-2,5-disubstituted pyrrolidine backbone were synthesized from commercially available starting materials meso-diethyl-2,5-dibromoadipate, (S)-(−)-1-phenylethylamine, and phenylalanine. The configurations of all the chiral centers in these unnatural dipeptides are confirmed by X-ray crystal diffraction analysis.

Keywords: cis-2,5-Disubstituted pyrrolidine; Unnatural dipeptide; Phenylalanine; X-ray diffraction analysis

Pharmacokinetics and cerebral distribution of glycine administered to rats by Nobuhiro Kawai; Makoto Bannai; Shinobu Seki; Tomonori Koizumi; Kenji Shinkai; Kenji Nagao; Daisuke Matsuzawa; Michio Takahashi; Eiji Shimizu (pp. 2129-2137).

High doses of glycine have been reported to improve negative schizophrenic symptoms, suggesting that ingested glycine activates glutamatergic transmission via N-methyl-d-aspartate (NMDA) receptors. However, the pharmacokinetics of administered glycine in the brain has not been evaluated. In the present study, the time- and dose-dependent distributions of administered glycine were investigated from a pharmacokinetic viewpoint. Whole-body autoradiography of radiolabeled glycine was performed, and time–concentration curves for glycine and serine in plasma, cerebrospinal fluid (CSF), and brain tissues were obtained. Furthermore, pharmacokinetic parameters were calculated. For a more detailed analysis, the amount of glycine uptake in the brain was evaluated using the brain uptake index method. Radiolabeled glycine was distributed among periventricular organs in the brain. Oral administration of 2 g/kg of glycine significantly elevated the CSF glycine concentration above the ED₅₀ value for NMDA receptors. The glycine levels in CSF were 100 times lower than those in plasma. Glycine levels were elevated in brain tissue, but with a slower time-course than in CSF. Serine, a major metabolite of glycine, was elevated in plasma, CSF, and brain tissue. Glycine uptake in brain tissue increased in a dose-dependent manner. Time–concentration curves revealed that glycine was most likely transported via the blood–CSF barrier and activated NMDA receptors adjacent to the ventricles. The pharmacokinetic analysis and the brain uptake index for glycine suggested that glycine was transported into brain tissue by passive diffusion. These results provide further insight into the potential therapeutic applications of glycine.

Keywords: Brain uptake index; Glycine; Schizophrenia; NMDA receptor; Blood–CSF barrier; Serine

Taurine-like GABA aminotransferase inhibitors prevent rabbit brain slices against oxygen–glucose deprivation-induced damage by Lorenzo Ricci; Massimo Valoti; Giampietro Sgaragli; Maria Frosini (pp. 2139-2147).

The activation of the GABAergic system has been shown to protect brain tissues against the damage that occurs after cerebral ischaemia. On the other hand, the taurine analogues (±)Piperidine-3-sulphonic- (PSA), 2-aminoethane phosphonic- (AEP), 2-(N-acetylamino) cyclohexane sulfonic-acids (ATAHS) and 2-aminobenzene sulfonate-acids (ANSA) have been reported to block GABA metabolism by inhibiting rabbit brain GABA aminotransferase and to increase GABA content in rabbit brain slices. The present investigation explored the neuroprotection provided by GABA, Vigabatrin (VIGA) and taurine analogues in the course of oxygen–glucose deprivation and reperfusion induced damage of rabbit brain slices. Tissue damage was assessed by measuring the release of glutamate and lactate dehydrogenase (LDH) during reperfusion and by determining final tissue water gain, measured as the index of cell swelling. GABA (30–300 μM) and VIGA (30–300 μM) significantly antagonised LDH and glutamate release, as well as tissue water gain caused by oxygen–glucose deprivation and reperfusion. Lower (1–10 μM) or higher concentrations (up to 3,000 μM) were ineffective. ANSA, PSA and ATAHS significantly reduced glutamate and LDH release and tissue water gain in a range of concentrations between 30 and 300 μM. Lower (0–10 μM) or higher (up to 3,000 μM) concentrations were ineffective. Both mechanisms suggest hormetic (“U-shaped”) effects. These results indicate that the GABAergic system activation performed directly by GABA or indirectly through GABA aminotransferase inhibition is a promising approach for protecting the brain against ischemia and reperfusion-induced damage.

Keywords: Taurine analogues; GABA; GABA aminotransferase inhibitors; Brain ischemia; Neuroprotection; Oxygen–glucose deprivation

Identification of c-myc-dependent proteins in the medulloblastoma cell line D425Med by Amedeo A. Azizi; Lin Li; Thomas Ströbel; Wei-Qiang Chen; Irene Slavc; Gert Lubec (pp. 2149-2163).

High c-myc levels are linked to poor prognosis in medulloblastoma (MB), and it was the aim of the current study to search for c-myc-dependent proteins in the MB cell line D425Med. For this purpose D425Med cells and cells with knocked-down c-myc (by siRNA) were analysed by a gel-based differential proteomics study using mass spectrometry. Heterogeneous nuclear ribonucleoproteins C1/C2, heterogeneous nuclear ribonucleoprotein A/B, stathmin, endoplasmic reticulum protein ERp29 precursor and guanidinoacetate N-methyltransferase were c-myc dependently expressed. Signalling, the protein machinery, metabolism and endoplasmic reticulum function may be affected and these results enable studying tumour tissue for these proteins as potential dignity markers or pharmacological targets.

Keywords: Brain tumour; Medulloblastoma; c-myc; myc

Decreasing the configurational entropy and the hydrophobicity of EBV-derived peptide 11389 increased its antigenicity, immunogenicity and its ability of inducing IL-6 by Mauricio Urquiza; Tatiana Guevara; Cristina Rodriguez; Johanna Melo-Cardenas; Magnolia Vanegas; Manuel E. Patarroyo (pp. 2165-2175).

Peptide 11389 from CD21-binding region of EBV-gp350/220 protein binds to PBMCs inducing IL-6 expression and inhibiting EBV-binding to PBMCs. In addition, anti-peptide 11389 antibodies recognize EBV-infected cells and inhibit both EBV infection and IL-6 production in PBMCs. We have postulated that native structure stabilization of peptide 11389 sequence can increase its biological activity. The strategy was to modify its sequence to restrict the number of structures that peptide 11389 could acquire in solution (decreasing peptide’s configurational entropy) and to weaken the non-relevant intermolecular interactions (decreasing its hydrophobicity), preserving CD21-interacting residues and structure as displayed in the native protein. Thirteen analog peptides were designed and synthesized; most of them were monomers containing an intra-chain disulfide bridge. Analog peptides 34058, 34060, 34061, 34296, 34298, 34299 and 34300 inhibited EBV invasion of PBMCs. Peptides 34059, 34060, 34295 and 34297 induced IL-6 levels in PBMCs (EC50 = 3.4, 3.3, 0.5, 0.5 μM, respectively) at higher potency than peptide 11389 (EC50 = 5.8 μM). Peptides 34057, 34059, 34060, 34301 and 34302 interacted with anti-EBV antibodies with affinities from 3 to 50 times higher than peptide 11389. Most of analog peptides were highly immunogenic and elicited antibodies that cross-react with EBV. In conclusion, we have designed peptides displaying higher biological activity than peptide 11389.

Keywords: EBV; Peptides; Entropy; IL-6; gp350; Antibodies

Rapid synthesis of new block copolyurethanes derived from l-leucine cyclodipeptide in reusable molten ammonium salts: novel and efficient green media for the synthesis of new hydrolysable and biodegradable copolyurethanes by Fatemeh Rafiemanzelat; Elahe Abdollahi (pp. 2177-2186).

This study concerns the synthesis of novel multi block polyurethane (PU) copolymers containing cyclodipeptide, taking the advantage of ionic liquids (ILs) under microwave irradiation. For this, l-leucine anhydride cyclodipeptide (LACP) was prepared and then a new class of poly(ether-urethane-urea)s (PEUUs) was synthesized in molten ammonium type ILs. ILs were used as reaction media and PUs were prepared via two-step polymerization method. In the first step, 4,4′-methylene-bis-(4-phenylisocyanate) (MDI) was reacted with LACP to produce isocyanate-terminated oligo(imide-urea) as hard segment (NCO-OIU). Chain extension of the aforementioned pre-polymer with polyethyleneglycol (PEG) of molecular weights of 1000 (PEG-1000) was the second step to furnish a series of new PEUUs. These multiblock copolymers are thermally stable, soluble in amide-type solvents, hydrolysable and biodegradable. PEUUs prepared in ILs under microwave irradiation showed more phase separation and crystallinity than PEUUs prepared under conventional method. The protocol presented here has the merits of environmentally benign, simple operation, convenient work-up, short reaction time and good yields without using volatile organic solvents, and catalysts. Ammonium type reaction media were air and water stable, and relatively cheap, which makes them suitable for application. The results demonstrate that they can be easily separated into water and reused without losing activity. Reusability of tetrabutylammonium bromide as reaction media makes the method a cost effective and environmentally benign method under microwave irradiation. Thus, we could prepare environmentally friendly polymers via environmentally benign method.

Keywords: Amino acid cyclodipeptide; Ionic liquids; Thermally stable polymers; Poly(ether-urethane-urea)s; Microwave assisted copolymerization; Green chemistry

Synthesis and properties of optically active nanostructured polymers bearing amino acid moieties by direct polycondensation of 4,4′-thiobis(2-tert-butyl-5-methylphenol) with chiral diacids by Shadpour Mallakpour; Samaneh Soltanian (pp. 2187-2194).

Four derivatives of N-trimellitylimido-l-amino acid (4a–4d) were prepared by the reaction of trimellitic anhydride (1) with the l-amino acids (2a–2d) in acetic acid as diacid monomers and were used with the aim to obtain a new family of amino acid based poly(ester-imide)s (PEI)s. The polymerization was performed by direct polycondensation of chiral diacids (4a–4d) with 4,4′-thiobis(2-tert-butyl-5-methylphenol) (5) in the presence of tosyl chloride (TsCl), pyridine and N,N-dimethyl formamide (DMF). Step-growth polymerization was carried out by varying the time of heating and the molar ratio of TsCl/diacid and the optimum conditions were achieved. The synthesized polymers were characterized by means of specific rotation experiments, FT-IR, ¹H-NMR, X-ray diffraction techniques and elemental analysis. The surface morphology of the obtained polymers was studied by field emission scanning electron microscopy. The result showed nanostructure morphology of the resulting polymers. The obtained PEIs were soluble in polar aprotic solvents such as DMF, N,N-dimethyl acetamide, dimethyl sulfoxide, N-methyl-2-pyrrolidone and protic solvents such as sulfuric acid. Thermal stability and the weight-loss behavior of the PEIs were studied by thermal gravimetric analysis (TGA) techniques. TGA showed that the 10% weight loss temperature in a nitrogen atmosphere was more than 402°C, therefore they had useful levels of thermal stability associated with excellent solubility.

Keywords: Poly(ester-imide); Nanostructure; Chiral polymers; l-Amino acid; Biodegradable

SOMPNN: an efficient non-parametric model for predicting transmembrane helices by Dong-Jun Yu; Hong-Bin Shen; Jing-Yu Yang (pp. 2195-2205).

Accurately predicting the transmembrane helices (TMH) in a helical membrane protein is an important but challenging task. Recent researches have demonstrated that statistics-based methods are promising routes to improve the TMH prediction accuracy. However, most of existing TMH predictors are parametric models and they have to make assumptions of several or even hundreds of adjustable parameters based on the underlying probability distribution, which is difficult when no a priori knowledge is available. Besides the performances of these parametric predictors significantly depend on the estimated parameters, some of them need to exploit the entire training dataset in the prediction stage, which will lead to low prediction efficiency and this problem will become even worse when dealing with large-scale dataset. In this paper, we propose a novel SOMPNN model for prediction of TMH that features by minimal parameter assumptions requirement and high computational efficiency. In the SOMPNN model, a self-organizing map (SOM) is used to adaptively learn the helices distribution knowledge hidden in the training data, and then a probabilistic neural network (PNN) is adopted to predict TMH segments based on the knowledge learned by SOM. Experimental results on two benchmark datasets show that the proposed SOMPNN outperforms most existing popular TMH predictors and is promising to be extended to deal with other complicated biological problems. The datasets and the source codes of SOMPNN are available at http://www.csbio.sjtu.edu.cn/bioinf/SOMPNN/ .

Keywords: Membrane protein; Transmembrane helix prediction; Non-parametric model; Self-organizing map; Probabilistic neural network; SOMPNN

Dietary l-arginine supplementation enhances placental growth and reproductive performance in sows by Kaiguo Gao; Zongyong Jiang; Yingcai Lin; Chuntian Zheng; Guilian Zhou; Fang Chen; Lin Yang; Guoyao Wu (pp. 2207-2214).

Suboptimal embryonic/fetal survival and growth remains a significant problem in mammals. Using a swine model, we tested the hypothesis that dietary l-arginine supplementation during gestation may improve pregnancy outcomes through enhancing placental growth and modulating hormonal secretions. Gestating pigs (Yorkshire × Landrace, n = 108) were assigned randomly into two groups based on parity and body weight, representing dietary supplementation with 1.0% l-arginine–HCl or 1.7% l-alanine (isonitrogenous control) between days 22 and 114 of gestation. Blood samples were obtained from the ear vein on days 22, 40, 70 and 90 of gestation. On days 40, 70 and 90 of gestation, concentrations of estradiol in plasma were higher (P < 0.05) in arginine-supplemented than in control sows. Moreover, arginine supplementation increased (P < 0.05) the concentrations of arginine, proline and ornithine in plasma, but concentrations of urea or progesterone in plasma did not differ between the two groups of sows. Compared with the control, arginine supplementation increased (P < 0.05) the total number of piglets by 1.31 per litter, the number of live-born piglets by 1.10 per litter, the litter birth weight for all piglets by 1.36 kg, and the litter birth weight for live-born piglets by 1.70 kg. Furthermore, arginine supplementation enhanced (P < 0.05) placental weight by 16.2%. The weaning-to-estrus interval of sows was not affected by arginine supplementation during gestation. These results indicate that dietary arginine supplementation beneficially enhances placental growth and the reproductive performance of sows.

Keywords: l-Arginine; Sow; Placenta; Estradiol; Reproductive performance; Piglet

A bursal pentapeptide (BPP-I), a novel bursal-derived peptide, exhibits antiproliferation of tumor cell and immunomodulator activity by Xiu L. Feng; Qing T. Liu; Rui B. Cao; Bin Zhou; Fang Q. Wang; Wen L. Deng; Ya F. Qiu; Yu Zhang; Hassan Ishag; Zhi Y. Ma; Qi S. Zheng; Pu Y. Chen (pp. 2215-2222).

The bursa of Fabricius (BF) is the central humoral immune organ unique to birds. Here, we isolated a novel bursal pentapeptide I (BPP-I), LGPGP, from BF. BPP-I could play inhibition effect on MCF-7 but not on CEF or Vero cell proliferation in vitro, and enhance antitumor factor p53 protein expression. Also, BPP-I stimulated antibody production in a dose-dependent manner in hybridoma cell. Furthermore, BPP-I could induce various immune responses in mice immunization experiments, including increase antibody production and cytokines IL-4 and IFN-γ level, and induce T-cell immunophenotyping. These results suggest that BPP-I is a potential immunomodulator of antitumor and immunity. The study could provide some novel insights on the probable candidate reagent for the antitumor and immune improvement.

Keywords: Bursal pentapeptide; Antiproliferation on tumor cell; Antitumor factor p53; Hybridoma cell; Immunodominant functions

Mechanism underlying the antioxidant activity of taurine: prevention of mitochondrial oxidant production by Chian Ju Jong; Junichi Azuma; Stephen Schaffer (pp. 2223-2232).

An important function of the β-amino acid, taurine, is the regulation of oxidative stress. However, taurine is neither a classical scavenger nor a regulator of the antioxidative defenses, leaving uncertain the mechanism underlying the antioxidant activity of taurine. In the present study, the taurine antagonist and taurine transport inhibitor, β-alanine, was used to examine the mechanism underlying the antioxidant activity of taurine. Exposure of isolated cardiomyocytes to medium containing β-alanine for a period of 48 h led to a 45% decrease in taurine content and an increase in mitochondrial oxidative stress, as evidenced by enhanced superoxide generation, the inactivation of the oxidant sensitive enzyme, aconitase, and the oxidation of glutathione. Associated with the increase in oxidative stress was a decline in electron transport activity, with the activities of respiratory chain complexes I and III declining 50–65% and oxygen consumption falling 30%. A reduction in respiratory chain activity coupled with an increase in oxidative stress is commonly caused by the development of a bottleneck in electron transport that leads to the diversion of electrons from the respiratory chain to the acceptor oxygen forming in the process superoxide. Because β-alanine exposure significantly reduces the levels of respiratory chain complex subunits, ND5 and ND6, the bottleneck in electron transport appears to be caused by impaired synthesis of key subunits of the electron transport chain complexes. Co-administration of taurine with β-alanine largely prevents the mitochondrial effects of β-alanine, but treatment of the cells with 5 mM taurine in the absence of β-alanine has no effect on the mitochondria, likely because taurine treatment has little effect on cellular taurine levels. Thus, taurine serves as a regulator of mitochondrial protein synthesis, thereby enhancing electron transport chain activity and protecting the mitochondria against excessive superoxide generation.

Keywords: Taurine; β-Alanine; Mitochondrial protein synthesis; Oxidative stress; Electron transport

Effects of the regulatory ligands calcium and GTP on the thermal stability of tissue transglutaminase by Carlo Cervellati; Katy Montin; Monica Squerzanti; Carlo Mischiati; Carlo Ferrari; Francesco Spinozzi; Paolo Mariani; Heinz Amenitsch; Carlo M. Bergamini; Vincenzo Lanzara (pp. 2233-2242).

Tissue transglutaminase undergoes thermal inactivation with first-order kinetics at moderate temperatures, in a process which is affected in opposite way by the regulatory ligands calcium and GTP, which stabilize different conformations. We have explored the processes of inactivation and of unfolding of transglutaminase and the effects of ligands thereon, combining approaches of differential scanning calorimetry (DSC) and of thermal analysis coupled to fluorescence spectroscopy and small angle scattering. At low temperature (38–45°C), calcium promotes and GTP protects from inactivation, which occurs without detectable disruption of the protein structure but only local perturbations at the active site. Only at higher temperatures (52–56°C), the protein structure undergoes major rearrangements with alterations in the interactions between the N- and C-terminal domain pairs. Experiments by DSC and fluorescence spectroscopy clearly indicate reinforced and weakened interactions of the domains in the presence of GTP and of calcium, and different patterns of unfolding. Small angle scattering experiments confirm different pathways of unfolding, with attainment of limiting values of gyration radius of 52, 60 and 90 Å in the absence of ligands and in the presence of GTP and calcium. Data by X-rays scattering indicate that ligands influence retention of a relatively compact structure in the protein even after denaturation at 70°C. These results suggest that the complex regulation of the enzyme by ligands involves both short- and long-range effects which might be relevant for understanding the turnover of the protein in vivo.

Keywords: Tissue transglutaminase; Regulatory ligands; Thermal inactivation; Fluorescence spectroscopy; Differential scanning calorimetry; Small-angle scattering

Accurate prediction of protein structural class using auto covariance transformation of PSI-BLAST profiles by Taigang Liu; Xingbo Geng; Xiaoqi Zheng; Rensuo Li; Jun Wang (pp. 2243-2249).

Computational prediction of protein structural class based solely on sequence data remains a challenging problem in protein science. Existing methods differ in the protein sequence representation models and prediction engines adopted. In this study, a powerful feature extraction method, which combines position-specific score matrix (PSSM) with auto covariance (AC) transformation, is introduced. Thus, a sample protein is represented by a series of discrete components, which could partially incorporate the long-range sequence order information and evolutionary information reflected from the PSI-BLAST profile. To verify the performance of our method, jackknife cross-validation tests are performed on four widely used benchmark datasets. Comparison of our results with existing methods shows that our method provides the state-of-the-art performance for structural class prediction. A Web server that implements the proposed method is freely available at http://202.194.133.5/xinxi/AAC_PSSM_AC/index.htm .

Keywords: Protein structural class; PSI-BLAST profile; Auto covariance transformation; Support vector machine

p70 S6K1 nuclear localization depends on its mTOR-mediated phosphorylation at T389, but not on its kinase activity towards S6 by M. Rosner; K. Schipany; M. Hengstschläger (pp. 2251-2256).

The protein kinase p70 S6K1 is regulated in response to cytokines, nutrients and growth factors, and plays an important role in the development of a variety of human diseases. Mammalian target of rapamycin (mTOR) is known to phosphorylate and thereby activate p70 S6K1. p70 S6K1 phosphorylates different cytoplasmic and nuclear substrates involved in the regulation of protein synthesis, cell cycle, cell growth and survival. Recently, we have shown that mTOR-mediated phosphorylation of p70 S6K1 at T389 also regulates its nucleocytoplasmic localization. Since this phosphorylation is associated with its kinase activity the question whether p70 S6K1 phosphorylation or kinase activity is essential for its proper localization remained elusive. Recently, the chemical compound PF-4708671 has been demonstrated to block p70 S6K1 kinase activity while inducing its phosphorylation at T389. This potential of PF-4708671 to separate p70 S6K1 activity from its T389 phosphorylation allowed us to demonstrate that the proper nucleocytoplasmic localization of this kinase depends on its mTOR-mediated phosphorylation but not on its kinase activity. These findings provide important insights into the regulation of p70 S6K1 and allow a more detailed understanding of subcellular enzyme localization processes.

Keywords: Kinase; mTOR; p70 S6Kinase; Phosphorylation; S6

Identification of a novel HLA-A2-restricted cytotoxic T lymphocyte epitope from cancer-testis antigen PLAC1 in breast cancer by Wei Liu; Mingxia Zhai; Zongyin Wu; Yuanming Qi; Yahong Wu; Chao Dai; Meng Sun; Lu Li; Yanfeng Gao (pp. 2257-2265).

Identification of cytotoxic T lymphocyte (CTL) epitopes from tumor antigens is essential for the development of peptide vaccines against tumor immunotherapy. Among all the tumor antigens, the caner-testis (CT) antigens are the most widely studied and promising targets. PLAC1 (placenta-specific 1, CT92) was considered as a novel member of caner-testis antigen, which expressed in a wide range of human malignancies, most frequently in breast cancer. In this study, three native peptides and their analogues derived from PLAC1 were predicted by T cell epitope prediction programs including SYFPEITHI, BIMAS and NetCTL 1.2. Binding affinity and stability assays in T2 cells showed that two native peptides, p28 and p31, and their analogues (p28-1Y9 V, p31-1Y2L) had more potent binding activity towards HLA-A*0201 molecule. In ELISPOT assay, the CTLs induced by these four peptides could release IFN-γ. The CTLs induced by these four peptides from the peripheral blood mononuclear cells (PBMCs) of HLA-A*02⁺ healthy donor could lyse MCF-7 breast cancer cells (HLA-A*0201⁺, PLAC1⁺) in vitro. When immunized in HLA-A2.1/K^b transgenic mice, the peptide p28 could induce the most potent peptide-specific CTLs among these peptides. Therefore, our results indicated that the peptide p28 (VLCSIDWFM) could serve as a novel candidate epitope for the development of peptide vaccines against PLAC1-positive breast cancer.

Keywords: Breast cancer; PLAC1; Cytotoxic T lymphocyte; Epitope; Immunotherapy

Taurine transporter gene expression in peripheral mononuclear blood cells of type 2 diabetic patients by Loria Bianchi; Riccardo Lari; Roberto Anichini; Alessandra De Bellis; Angela Berti; Zaleida Napoli; Giuseppe Seghieri; Flavia Franconi (pp. 2267-2274).

Taurine acts as antioxidant, cell osmolyte, modulator of glucose metabolism, and plays a role in the retinal function. It is 10³-fold more concentrated in the intracellular than in the extracellular milieu due to a specific taurine-Na-dependent transporter (TauT), which is upregulated by hypertonicity, low extracellular taurine, or oxidative stress and acutely downregulated ‘in vitro’ by high glucose concentrations. Aim of this study was to investigate whether TauT expression was modified in mononuclear peripheral blood cells (MPC) of type 2 diabetic patients with or without micro/macrovascular complications. Plasma taurine, as well as other sulphur-containing aminoacids (assayed by HPLC) and TauT gene expression (assayed by real-time PCR analysis) were measured in MPC of 45 controls and of 81 age-and-sex matched type 2 diabetic patients with or without micro/macrovascular complications. Median value (interquartile range) of plasma taurine was significantly lower in diabetic patients than in controls [28.7 (13.7) μmol/l vs. 46.5 (20.3) μmol/l; P < 0.05], while median TauT expression, in arbitrary units, was significantly higher in diabetics than in controls [3.8 (3.9) vs. 1 (1.3); P < 0.05) and was related to HbA1c only in controls (r = 0.34; P < 0.05). Patients with retinopathy (n = 25) had lower TauT expression than those who were unaffected [3.1 (2.8) vs. 4.1 (3.4); P < 0.05], while persistent micro/macroalbuminuria was associated with unchanged TauT expression. A trend toward reduction in TauT expression was observed in patients with macroangiopathy [n = 27; 3.3 (2.5) vs. 4 [3.7]; P = NS]. In conclusion, TauT gene is overexpressed in MPC of type 2 diabetic patients, while presence of retinopathy is specifically associated with a drop in TauT overexpression, suggesting its possible involvement in this microangiopathic lesion.

Keywords: Taurine; Taurine transporter; Type 2 diabetes; Retinopathy; Mononuclear peripheral blood cells

Photorelease of amino acids from novel thioxobenzo[f]benzopyran ester conjugates by Ana M. Piloto; Ana M. S. Soares; Susana P. G. Costa; M. Sameiro T. Gonçalves (pp. 2275-2282).

Aiming at the enhancement of the performance of (9-methoxy-3-oxo-3H-benzo[f]benzopyran-1-yl) methyl ester as photocleavable protecting group for the carboxylic acid function at long-wavelengths, 9-methoxy-3-thioxo-3H-benzo[f]benzopyran-l-valine and l-phenylalanine model conjugates were prepared through a thionation reaction of the corresponding oxo-benzobenzopyrans. These thioxobenzobenzopyran derivatives were subjected to photocleavage reactions in the same conditions as the parent oxo-benzobenzopyrans at different wavelengths of irradiation, and photocleavage data were obtained. It was found that the exchange of the carbonyl by a thiocarbonyl group enhanced the performance of the heterocyclic protecting group at 419 nm by improving the photolysis rates, making it an appropriate group for practical applications at long wavelengths.

Keywords: Thioxobenzobenzopyran; Benzocoumarin; Amino acids; Photolabile protecting groups

δ¹-Pyrroline-5-carboxylate reductase as a new target for therapeutics: inhibition of the enzyme from Streptococcus pyogenes and effects in vivo by Giuseppe Forlani; Davide Petrollino; Massimo Fusetti; Letizia Romanini; Bogusław Nocek; Andrzej Joachimiak; Łukasz Berlicki; Paweł Kafarski (pp. 2283-2291).

Compounds able to interfere with amino acid biosynthesis have the potential to inhibit cell growth. In both prokaryotic and eukaryotic microorganisms, unless an ornithine cyclodeaminase is present, the activity of δ¹-pyrroline-5-carboxylate (P5C) reductase is mandatory to proline production, and the enzyme inhibition should result in amino acid starvation, blocking in turn protein synthesis. The ability of some substituted derivatives of aminomethylenebisphosphonic acid and its analogues to interfere with the activity of the enzyme from the human pathogen Streptococcus pyogenes was investigated. Several compounds were able to suppress activity in the micromolar range of concentrations, with a mechanism of uncompetitive type with respect to the substrate P5C and non-competitive with respect to the electron donor NAD(P)H. The actual occurrence of enzyme inhibition in vivo was supported by the effects of the most active derivatives upon bacterial growth and free amino acid content.

Keywords: Amino acid metabolism; Antibiotics; P5C reductase; Proline; Streptococcus sp

d-Ser-containing humanin shows promotion of fibril formation by Kanehiro Hayashi; Jumpei Sasabe; Tomohiro Chiba; Sadakazu Aiso; Naoko Utsunomiya-Tate (pp. 2293-2297).

Humanin (HN), a peptide of 24 amino acid residues, suppresses the neuronal cell death that is induced by the gene products of Alzheimer’s disease. HN contains two Ser residues at positions 7 and 14. Because the proportion of d-Ser isomerized from l-Ser in proteins appears to increase as cellular organs age, we explored the structural effects of the isomerization of each Ser residue in HN. By using a thioflavin-T assay to detect fibril formation, we found that an HN derivative that contained two isomerized d-Ser residues had a greater tendency to form fibrils than did wild-type HN or HNs containing single d-Ser residues. A previous report showed that HN containing two d-Ser residues exerts neuroprotective activity. Our data, therefore, suggest that the fibril formation by HN that contains two d-Ser residues may promote HN neuroprotective activity.

Keywords: Humanin; d-Ser; Circular dichroism; β-Sheet; α-Helix

Creatine but not betaine supplementation increases muscle phosphorylcreatine content and strength performance by Serena del Favero; Hamilton Roschel; Guilherme Artioli; Carlos Ugrinowitsch; Valmor Tricoli; André Costa; Renato Barroso; Ana Lua Negrelli; Maria Concepción Otaduy; Cláudia da Costa Leite; Antonio Herbert Lancha-Junior; Bruno Gualano (pp. 2299-2305).

We aimed to investigate the role of betaine supplementation on muscle phosphorylcreatine (PCr) content and strength performance in untrained subjects. Additionally, we compared the ergogenic and physiological responses to betaine versus creatine supplementation. Finally, we also tested the possible additive effects of creatine and betaine supplementation. This was a double-blind, randomized, placebo-controlled study. Subjects were assigned to receive betaine (BET; 2 g/day), creatine (CR; 20 g/day), betaine plus creatine (BET + CR; 2 + 20 g/day, respectively) or placebo (PL). At baseline and after 10 days of supplementation, we assessed muscle strength and power, muscle PCr content, and body composition. The CR and BET + CR groups presented greater increase in muscle PCr content than PL (p = 0.004 and p = 0.006, respectively). PCr content was comparable between BET versus PL (p = 0.78) and CR versus BET + CR (p = 0.99). CR and BET + CR presented greater muscle power output than PL in the squat exercise following supplementation (p = 0.003 and p = 0.041, respectively). Similarly, bench press average power was significantly greater for the CR-supplemented groups. CR and BET + CR groups also showed significant pre- to post-test increase in 1-RM squat and bench press (CR: p = 0.027 and p < 0.0001; BET + CR: p = 0.03 and p < 0.0001 for upper- and lower-body assessments, respectively) No significant differences for 1-RM strength and power were observed between BET versus PL and CR versus BET + CR. Body composition did not differ between the groups. In conclusion, we reported that betaine supplementation does not augment muscle PCr content. Furthermore, we showed that betaine supplementation combined or not with creatine supplementation does not affect strength and power performance in untrained subjects.

Keywords: Betaine supplementation; Creatine supplementation; Maximal muscle strength; Muscle power output; Phosphorylcreatine content

Arabidopsis mutants lacking asparaginases develop normally but exhibit enhanced root inhibition by exogenous asparagine by Ana Ivanov; Alexander Kameka; Agnieszka Pajak; Luanne Bruneau; Ronald Beyaert; Cinta Hernández-Sebastià; Frédéric Marsolais (pp. 2307-2318).

Asparaginase catalyzes the degradation of l-asparagine to l-aspartic acid and ammonia, and is implicated in the catabolism of transported asparagine in sink tissues of higher plants. The Arabidopsis genome includes two genes, ASPGA1 and ASPGB1, belonging to distinct asparaginase subfamilies. Conditions of severe nitrogen limitation resulted in a slight decrease in seed size in wild-type Arabidopsis. However, this response was not observed in a homozygous T-DNA insertion mutant where ASPG genes had been inactivated. Under nitrogen-sufficient conditions, the ASPG mutant had elevated levels of free asparagine in mature seed. This phenotype was observed exclusively under conditions of low illumination, when a low ratio of carbon to nitrogen was translocated to the seed. Mutants deficient in one or both asparaginases were more sensitive than wild-type to inhibition of primary root elongation and root hair emergence by l-asparagine as a single nitrogen source. This enhanced inhibition was associated with increased accumulation of asparagine in the root of the double aspga1-1/-b1-1 mutant. This indicates that inhibition of root growth is likely elicited by asparagine itself or an asparagine-derived metabolite, other than the products of asparaginase, aspartic acid or ammonia. During germination, a fusion between the ASPGA1 promoter and beta-glucuronidase was expressed in endosperm cells starting at the micropylar end. Expression was initially high throughout the root and hypocotyl, but became restricted to the root tip after three days, which may indicate a transition to nitrogen-heterotrophic growth.

Keywords: Asparaginase; Asparagine; Mutants; Root elongation; Root hair formation; Nutrient sensing

A short review on creatine–creatine kinase system in relation to cancer and some experimental results on creatine as adjuvant in cancer therapy by Subrata Patra; Alok Ghosh; Soumya Sinha Roy; Soumen Bera; Manju Das; Dipa Talukdar; Subhankar Ray; Theo Wallimann; Manju Ray (pp. 2319-2330).

The creatine/creatine kinase (CK) system plays a key role in cellular energy buffering and transport. In vertebrates, CK has four isoforms expressed in a tissue-specific manner. In the process of creatine biosynthesis several other important metabolites are formed. The anticancer effect of creatine had been reported in the past, and recent literature has reported low creatine content in several types of malignant cells. Furthermore, creatine can protect cardiac mitochondria from the deleterious effects of some anticancer compounds. Previous work from our laboratory showed progressive decrease of phosphocreatine, creatine and CK upon transformation of skeletal muscle into sarcoma. It was convincingly demonstrated that prominent expression of creatine-synthesizing enzymes l-arginine: glycine amidinotransferase and N-guanidinoacetate methyltransferase occurs in sarcoma, Ehrlich ascites carcinoma and sarcoma 180 cells; whereas, both these enzymes are virtually undetectable in skeletal muscle. Creatine transporter also remained unaltered in malignant cells. The anticancer effect of methylglyoxal had been known for a long time. The present work shows that this anticancer effect of methylglyoxal is significantly augmented in presence of creatine. On creatine supplementation the effect of methylglyoxal plus ascorbic acid was further augmented and there was no visible sign of tumor. Moreover, creatine and CK, which were very low in sarcoma tissue, were significantly elevated with the concomitant regression of tumor.

Keywords: Ascorbic acid; Cancer; Creatine; Creatine kinase; Methylglyoxal

Activation of carboxyl group with cyanate: peptide bond formation from dicarboxylic acids by Grégoire Danger; Solenne Charlot; Laurent Boiteau; Robert Pascal (pp. 2331-2341).

The reaction of cyanate with C-terminal carboxyl groups of peptides in aqueous solution was considered as a potential pathway for the abiotic formation of peptide bonds under the condition of the primitive Earth. The catalytic effect of dicarboxylic acids on cyanate hydrolysis was definitely attributed to intramolecular nucleophilic catalysis by the observation of the ¹H-NMR signal of succinic anhydride when reacting succinic acid with KOCN in aqueous solution (pH 2.2–5.5). The formation of amide bonds was noticed when adding amino acids or amino acid derivatives into the solution. The reaction of N-acyl aspartic acid derivatives was observed to proceed similarly and the scope of the cyanate-promoted reaction was analyzed from the standpoint of prebiotic peptide formation. The role of cyanate in activating peptide C-terminus constitutes a proof of principle that intramolecular reactions of adducts of peptides C-terminal carboxyl groups with activating agents represent a pathway for peptide activation in aqueous solution, the relevance of which is discussed in connexion with the issue of the emergence of homochirality.

Keywords: Prebiotic chemistry; Peptide; Activation; Intramolecular catalysis; Neighbouring group assistance

Imaging tumor-induced sentinel lymph node lymphangiogenesis with LyP-1 peptide by Fan Zhang; Gang Niu; Xin Lin; Orit Jacobson; Ying Ma; Henry S. Eden; Yulong He; Guangming Lu; Xiaoyuan Chen (pp. 2343-2351).

Lymphangiogenesis in tumor-draining lymph nodes (LNs) starts before the onset of metastasis and is associated with metastasis to distant LNs and organs. In this study, we aimed to visualize tumor-induced lymphangiogenesis with a tumor lymphatics-specific peptide LyP-1. The LyP-1 peptide was labeled with a near-infrared fluorophore (Cy5.5) for optical imaging. At days 3, 7, 14 and 21 after subcutaneous 4T1 tumor inoculation, Cy5.5-LyP-1 was administered through the middle phalanges of the upper extremities of the tumor-bearing mice. At 45 min and 24 h postinjection, brachial LN fluorescence imaging was performed. Ex vivo fluorescence images were acquired for quantitative analysis of the fluorescence intensity. Tumor-induced lymphangiogenesis was confirmed by LYVE-1 immunostaining and increased size of tumor side brachial LNs. Cy5.5-LyP-1 staining in LNs co-localized with LYVE-1, suggesting lymphatics-specific binding of LyP-1 peptide. The brachial LNs were clearly visualized by optical imaging at both time points. The tumor side LNs showed significantly higher fluorescence intensities than the contralateral brachial LNs at days 7, 14, and 21, but not day 3 after tumor inoculation. At day 21 after tumor inoculation, the average signal of tumor-draining LNs was 78.0 ± 2.44, 24.3 ± 5.43, 25.6 ± 0.25 (×10³ photon/cm²/s) using Cy5.5-LyP-1, Cy5.5-LyP-1 with blocking, and Cy5.5 only, respectively. Tumor-draining brachial LNs showed extensive growth of lymphatic sinuses throughout the cortex and medulla. Use of LyP-1 based imaging probes with optical imaging offers a useful tool for the study of tumor-induced lymphangiogenesis. LyP-1 may serve as a marker of lymphangiogenesis useful in detecting “high risk” LNs before tumor metastasis and after micro-metastasis, as well as for screening potential anti-lymphatic therapies.

Keywords: Lymphangiogenesis; Lymph node; LyP-1 peptide; Optical imaging

Statistical energy potential: reduced representation of Dehouck–Gilis–Rooman function by selecting against decoy datasets by Wen-Wei Lu; Ri-Bo Huang; Yu-Tuo Wei; Jian-Zong Meng; Li-Qin Du; Qi-Shi Du (pp. 2353-2361).

Statistical effective energy function (SEEF) is derived from the statistical analysis of the database of known protein structures. Dehouck–Gilis–Rooman (DGR) group has recently created a new generation of SEEF in which the additivity of the energy terms was manifested by decomposing the total folding free energy into a sum of lower order terms. We have tried to optimize the potential function based on their work. By using decoy datasets as screening filter, and through modification of algorithms in calculation of accessible surface area and residue–residue interaction cutoff, four new combinations of the energy terms were found to be comparable to DGR potential in performance test. Most importantly, the term number was reduced from the original 30 terms to only 5 in our results, thereby substantially decreasing the computation time while the performance was not sacrificed. Our results further proved the additivity and manipulability of the DGR original energy function, and our new combination of the energy could be used in prediction of protein structures.

Keywords: Decoy dataset; Performance of energy function; PyTables; Protein structure prediction; Statistical effective energy function (SEEF)

Prediction of protein–protein interactions between Ralstonia solanacearum and Arabidopsis thaliana by Zhi-Gang Li; Fei He; Ziding Zhang; You-Liang Peng (pp. 2363-2371).

Ralstonia solanacearum is a devastating bacterial pathogen that has an unusually wide host range. R. solanacearum, together with Arabidopsis thaliana, has become a model system for studying the molecular basis of plant–pathogen interactions. Protein–protein interactions (PPIs) play a critical role in the infection process, and some PPIs can initiate a plant defense response. However, experimental investigations have rarely addressed such PPIs. Using two computational methods, the interolog and the domain-based methods, we predicted 3,074 potential PPIs between 119 R. solanacearum and 1,442 A. thaliana proteins. Interestingly, we found that the potential pathogen-targeted proteins are more important in the A. thaliana PPI network. To facilitate further studies, all predicted PPI data were compiled into a database server called PPIRA ( http://protein.cau.edu.cn/ppira/ ). We hope that our work will provide new insights for future research addressing the pathogenesis of R. solanacearum.

Keywords: Bioinformatics; Pathogenicity; Plant–pathogen interactions; Prediction; Protein–protein interaction

Identification of peptide sequences that target to the brain using in vivo phage display by Jingwei Li; Qizhi Zhang; Zhiqing Pang; Yuchen Wang; Qingfeng Liu; Liangran Guo; Xinguo Jiang (pp. 2373-2381).

Phage display technology could provide a rapid means for the discovery of novel peptides. To find peptide ligands specific for the brain vascular receptors, we performed a modified phage display method. Phages were recovered from mice brain parenchyma after administrated with a random 7-mer peptide library intravenously. A longer circulation time was arranged according to the biodistributive brain/blood ratios of phage particles. Following sequential rounds of isolation, a number of phages were sequenced and a peptide sequence (CTSTSAPYC, denoted as PepC7) was identified. Clone 7-1, which encodes PepC7, exhibited translocation efficiency about 41-fold higher than the random library phage. Immunofluorescence analysis revealed that Clone 7-1 had a significant superiority on transport efficiency into the brain compared with native M13 phage. Clone 7-1 was inhibited from homing to the brain in a dose-dependent fashion when cyclic peptides of the same sequence were present in a competition assay. Interestingly, the linear peptide (ATSTSAPYA, Pep7) and a scrambled control peptide PepSC7 (CSPATSYTC) did not compete with the phage at the same tested concentration (0.2–200 pg). Labeled by Cy5.5, PepC7 exhibited significant brain-targeting capability in in vivo optical imaging analysis. The cyclic conformation of PepC7 formed by disulfide bond, and the correct structure itself play a critical role in maintaining the selectivity and affinity for the brain. In conclusion, PepC7 is a promising brain-target motif never been reported before and it could be applied to targeted drug delivery into the brain.

Keywords: Phage display; Blood–brain barrier (BBB); Brain targeting; Peptide

The extended loop of the C-terminal carbohydrate-recognition domain of Manduca sexta immulectin-2 is important for ligand binding and functions by Xiu-Zhen Shi; Xiao-Qiang Yu (pp. 2383-2391).

Our previous research showed that immulectin-2 (IML-2), a C-type lectin from the tobacco hornworn, Manduca sexta, is a pattern recognition receptor (PRR) that can bind to pathogen-associated molecular patterns (PAMPs), such as lipopolysaccharide (LPS), peptidoglycan (PG) and β-1,3-glucan, and IML-2 plays an important role in cellular encapsulation, melanization, phagocytosis, and prophenoloxidase (proPO) activation. Unlike most mammalian C-type lectins that contain a single carbohydrate-recognition domain (CRD), IML-2 is composed of tandem CRDs, and the C-terminal CRD2 contains an extended loop, which is not present in most C-type CRDs. We hypothesize that the extended loop may participate in ligand binding, encapsulation, melanization, phagocytosis and/or proPO activation in M. sexta. To test this hypothesis, two deletion mutant proteins (IML-2^Δ220–244 and IML-2^Δ220–257), in which the extended loop of the CRD2 was partially or completely deleted, were expressed and purified. By comparing the characteristics of recombinant IML-2, IML-2^Δ220–244 and IML-2^Δ220–257, we found that deletion of the extended loop in CRD2 impaired the ability of IML-2 to bind microbial PAMPs and to stimulate proPO activation, indicating that the extended loop of IML-2 plays an important role in ligand binding and biological functions.

Keywords: Immulectin-2; Carbohydrate-recognition domain; Extended loop; Ligand binding; Encapsulation and melanization; Prophenoloxidase activation

The antibacterial peptide ABP-CM4: the current state of its production and applications by Jian Feng Li; Jie Zhang; Xing Zhou Xu; Yang Yang Han; Xian Wei Cui; Yu Qing Chen; Shuang Quan Zhang (pp. 2393-2402).

The increasing resistance of bacteria and fungi to currently available antibiotics is a major concern worldwide, leading to enormous efforts to develop new antibiotics with new modes of actions. Antibacterial peptide CM4 (ABP-CM4) is a small cationic peptide with broad-spectrum activities against bacteria, fungi, and tumor cells, which may possibly be used as a promising candidate for a new antibiotic. For pharmaceutical applications, a large quantity of antimicrobial peptides needs to be produced economically. In this communication, the progress in the structural characteristics, heterologous production, and biological evaluation of ABP-CM4 are reviewed.

Keywords: Antibacterial peptide; ABP-CM4; Heterologous expression; Antimicrobial; Anticancer

Taurine enhances the anorexigenic effects of insulin in the hypothalamus of rats by Carina S. Solon; Daniel Franci; Letícia M. Ignacio-Souza; Talita Romanatto; Erika A. Roman; Ana P. Arruda; Joseane Morari; Adriana S. Torsoni; Everardo M. Carneiro; Licio A. Velloso (pp. 2403-2410).

Taurine is known to modulate a number of metabolic parameters such as insulin secretion and action and blood cholesterol levels. Recent data have suggested that taurine can also reduce body adiposity in C. elegans and in rodents. Since body adiposity is mostly regulated by insulin-responsive hypothalamic neurons involved in the control of feeding and thermogenesis, we hypothesized that some of the activity of taurine in the control of body fat would be exerted through a direct action in the hypothalamus. Here, we show that the intracerebroventricular injection of an acute dose of taurine reduces food intake and locomotor activity, and activates signal transduction through the Akt/FOXO1, JAK2/STAT3 and mTOR/AMPK/ACC signaling pathways. These effects are accompanied by the modulation of expression of NPY. In addition, taurine can enhance the anorexigenic action of insulin. Thus, the aminoacid, taurine, exerts a potent anorexigenic action in the hypothalamus and enhances the effect of insulin on the control of food intake.

Keywords: Obesity; Leptin; Neurotransmitter

Carnosine treatment largely prevents alterations of renal carnosine metabolism in diabetic mice by Verena Peters; Claus P. Schmitt; Johannes Zschocke; Marie-Luise Gross; Kerstin Brismar; Elisabete Forsberg (pp. 2411-2416).

Recently, we identified an allelic variant of human carnosinase 1 (CN1) that results in increased enzyme activity and is associated with susceptibility for diabetic nephropathy in humans. Investigations in diabetic (db/db) mice showed that carnosine ameliorates glucose metabolism effectively. We now investigated the renal carnosinase metabolism in db/db mice. Kidney CN1 activity increased with age and was significantly higher in diabetic mice compared to controls. Increased CN1 activity did not affect renal carnosine levels, but anserine concentrations were tenfold lower in db/db mice compared to controls (0.24 ± 0.2 vs. 2.28 ± 0.3 nmol/mg protein in controls; p < 0.001). Homocarnosine concentrations in kidney tissue were low in both control and db/db mice (below 0.1 nmol/mg protein, p = n.s.). Carnosine treatment for 4 weeks substantially decreased renal CN1 activity in diabetic mice (0.32 ± 0.3 in non-treated db/db vs. 0.05 ± 0.05 μmol/mg/h in treated db/db mice; p < 0.01) close to normal activities. Renal anserine concentrations increased significantly (0.24 ± 0.2 in non-treated db/db vs. 5.7 ± 1.2 μmol/mg/h in treated db/db mice; p < 0.01), while carnosine concentrations remained unaltered (53 ± 6.4 in non-treated vs. 61 ± 15 nmol/mg protein in treated db/db mice; p = n.s.). Further, carnosine treatment halved proteinuria and reduced vascular permeability to one-fifth in db/db mice. In renal tissue of diabetic mice carnosinase activity is significantly increased and anserine concentrations are significantly reduced compared to controls. Carnosine treatment largely prevents the alterations of renal carnosine metabolism.

Keywords: Carnosine; Anserine; Diabetic mice (db/db); Carnosinase (CN1); Vascular permeability

Taurine attenuates Streptococcus uberis-induced mastitis in rats by increasing T regulatory cells by Jinfeng Miao; Jinqiu Zhang; Liuhai Zheng; Xiaoming Yu; Wei Zhu; Sixiang Zou (pp. 2417-2428).

Taurine (Tau) is reported to have a key role in the regulation of the innate immune response and thus reduce tissue damage induced by bacterial infection. In this study, the effects of Tau on a rat model of mastitis induced by Streptococcus uberis (S. uberis) and the changes of T regulatory cells (Tregs) were assessed. Starting on gestation day 14 and continuing until parturition, 100 mg/kg of taurine (group TS) or an equal volume of physiological saline (group CS) was administered daily, per os. Seventy-two hours after parturition, rats were infused with approximately 100 cfu of S. uberis into each of two mammary glands. The results showed that the resultant inflammation, evidenced by swelling, secretory epithelial cell degeneration, increased adipose tissue and neutrophil (PMN) infiltration were evident in mammary tissue following injection with S. uberis. Pre-treatment with Tau attenuated these morphologic changes, the expression of interleukin (IL)-2, interferon (INF)-γ mRNA, myeloperoxidase (MPO) activity and N-acetyl-β-D-glucosaminidase (NAGase) in mammary tissue. The percentages of Foxp3 + CD25 + CD4 +/lymphocytes (Tregs) were dramatically increased after the S. uberis challenge. Significant differences (P < 0.05) were observed at 24, and 72 h post S. uberis - injection (PI) in CS. Pre-treatment further increased the percentage of Tregs and a significant difference between CS and TS (P < 0.05) was apparent at 24 h PI. Our data indicate that in rats, Tau can be used to regulate the immune response following infection by S. uberis and consequently prevent mammary tissue damage by increasing Tregs.

Keywords: Taurine; Streptococcus uberis ; Mastitis; Rat; Tregs

Cross-linking of wheat gluten proteins during production of hard pretzels by Ine Rombouts; Bert Lagrain; Kristof Brijs; Jan A. Delcour (pp. 2429-2438).

The impact of the hot alkaline dip, prior to pretzel-baking, on the types and levels of cross-links between wheat proteins was studied. Protein extractability of pretzel dough in sodium dodecyl sulfate containing buffer decreased during alkaline dipping [45 s, 1.0% (w/v) NaOH, 90°C], and even more during baking (3 min at 250°C) and drying (10 min at 135°C). Reducing agent increased the extractability partly, indicating that both reducible (disulfide, SS) and non-reducible (non-SS) protein cross-links had been formed. The decrease in cystine levels suggested β-elimination of cystine releasing Cys and dehydroalanine (DHA). Subsequent reaction of DHA with Lys and Cys, induced the unusual and potentially cross-linking amino acids lysinoalanine (LAL) and lanthionine (LAN), respectively, in alkaline dipped dough (7 μmol LAN/g protein) and in the end product (9 μmol LAL and 50 μmol LAN/g protein). The baking/drying step increased sample redness, decreased Lys levels more than expected based on LAL formation (57 μmol/g protein), and induced a loss of reducing sugars (99 μmol/g protein), which suggested the potential contribution of Maillard-derived cross-links to the observed extractability loss. However, levels of Maillard products which possibly cross-link proteins, are small compared to DHA-derived cross-links. Higher dipping temperatures, longer dipping times, and higher NaOH concentrations increased protein extractability losses and redness, as well as LAL and LAN levels in the end product. No indications for Maillard-derived cross-links or LAL in pretzel dough immediately after dipping were found, even when severe dipping conditions were used.

Keywords: Gluten; Beta-elimination; Dehydroalanine; Lysinoalanine; Lanthionine

The reaction of methionine with hydroxyl radical: reactive intermediates and methanethiol production by Ivan Spasojević; Jelena Bogdanović Pristov; Ljubodrag Vujisić; Mihajlo Spasić (pp. 2439-2445).

The mechanisms of reaction of methionine with hydroxyl radical are not fully understood. Here, we unequivocally show using electron paramagnetic resonance spin-trapping spectroscopy and GC–FID and GC–MS, the presence of specific carbon-, nitrogen- and sulfur-centered radicals as intermediates of this reaction, as well as the liberation of methanethiol as a gaseous end product. Taking into account the many roles that methionine has in eco- and biosystems, our results may elucidate redox chemistry of this amino acid and processes that methionine is involved in.

Keywords: Methionine; Hydroxyl radical; EPR; GC; Free radical; Methanethiol

Prediction of membrane proteins using split amino acid and ensemble classification by Maqsood Hayat; Asifullah Khan; Mohammed Yeasin (pp. 2447-2460).

Knowledge of the types of membrane protein provides useful clues in deducing the functions of uncharacterized membrane proteins. An automatic method for efficiently identifying uncharacterized proteins is thus highly desirable. In this work, we have developed a novel method for predicting membrane protein types by exploiting the discrimination capability of the difference in amino acid composition at the N and C terminus through split amino acid composition (SAAC). We also show that the ensemble classification can better exploit this discriminating capability of SAAC. In this study, membrane protein types are classified using three feature extraction and several classification strategies. An ensemble classifier Mem-EnsSAAC is then developed using the best feature extraction strategy. Pseudo amino acid (PseAA) composition, discrete wavelet analysis (DWT), SAAC, and a hybrid model are employed for feature extraction. The nearest neighbor, probabilistic neural network, support vector machine, random forest, and Adaboost are used as individual classifiers. The predicted results of the individual learners are combined using genetic algorithm to form an ensemble classifier, Mem-EnsSAAC yielding an accuracy of 92.4 and 92.2% for the Jackknife and independent dataset test, respectively. Performance measures such as MCC, sensitivity, specificity, F-measure, and Q-statistics show that SAAC-based prediction yields significantly higher performance compared to PseAA- and DWT-based systems, and is also the best reported so far. The proposed Mem-EnsSAAC is able to predict the membrane protein types with high accuracy and consequently, can be very helpful in drug discovery. It can be accessed at http://111.68.99.218/membrane .

Keywords: Membrane protein types; Split amino acid; Discrete wavelet transform; Pseudo amino acid composition; Neural networks; Ensemble classifier; Random forest

Effect of two β-alanine dosing protocols on muscle carnosine synthesis and washout by Trent Stellingwerff; Helen Anwander; Andrea Egger; Tania Buehler; Roland Kreis; Jacques Decombaz; Chris Boesch (pp. 2461-2472).

Carnosine (β-alanyl-l-histidine) is found in high concentrations in skeletal muscle and chronic β-alanine (BA) supplementation can increase carnosine content. This placebo-controlled, double-blind study compared two different 8-week BA dosing regimens on the time course of muscle carnosine loading and 8-week washout, leading to a BA dose–response study with serial muscle carnosine assessments throughout. Thirty-one young males were randomized into three BA dosing groups: (1) high–low: 3.2 g BA/day for 4 weeks, followed by 1.6 g BA/day for 4 weeks; (2) low–low: 1.6 g BA/day for 8 weeks; and (3) placebo. Muscle carnosine in tibialis-anterior (TA) and gastrocnemius (GA) muscles was measured by ¹H-MRS at weeks 0, 2, 4, 8, 12 and 16. Flushing symptoms and blood clinical chemistry were trivial in all three groups and there were no muscle carnosine changes in the placebo group. During the first 4 weeks, the increase for high–low (TA 2.04 mmol/kg_ww, GA 1.75 mmol/kg_ww) was ~twofold greater than low–low (TA 1.12 mmol/kg_ww, GA 0.80 mmol/kg_ww). 1.6 g BA/day significantly increased muscle carnosine within 2 weeks and induced continual rises in already augmented muscle carnosine stores (week 4–8, high–low regime). The dose–response showed a carnosine increase of 2.01 mmol/kg_ww per 100 g of consumed BA, which was only dependent upon the total accumulated BA consumed (within a daily intake range of 1.6–3.2 g BA/day). Washout rates were gradual (0.18 mmol/kg_ww and 0.43 mmol/kg_ww/week; ~2%/week). In summary, the absolute increase in muscle carnosine is only dependent upon the total BA consumed and is not dependent upon baseline muscle carnosine, the muscle type, or the daily amount of supplemented BA.

Keywords: β-alanine; Carnosine; Muscle; Synthesis; Washout; Dose–response

The non-protein amino acid β-N-methylamino-l-alanine in Portuguese cyanobacterial isolates by Rosa C. Cervantes Cianca; Mafalda S. Baptista; Viviana R. Lopes; Vitor M. Vasconcelos (pp. 2473-2479).

The tailor made amino acid β-N-methyl-amino-l-alanine (BMAA) is a neurotoxin produced by cyanobacteria. It has been associated with certain forms of progressive neurodegenerative disease, including sporadic Amyotrophic Lateral Sclerosis and Alzheimer’s disease. Some different reports of BMAA in cyanobacterial blooms from lakes, reservoirs, and other water resources have been made by different investigators. We here report the detection of BMAA of both free and protein-bound produced by cyanobacteria, belonging to the Chroococcales, Oscillatoriales and Nostocales ordered. We use a rapid and sensitive HPLC-FD method that utilizes methanol elution and the Waters AQC Tag chemistry. On other hand, we have used three different assay procedures for BMAA extraction from cyanobacteria: Trichloroacetic acid (TCA), Methanol/Acetone and hydrochloric acid (HCl). All assays let successfully detect BMAA in all cyanobacteria samples analyzed. Nevertheless, with TCA and HCl extraction procedures the highest BMAA values, for free as well as protein-bound BMAA were detected. BMAA content could not be related to the taxonomy of the isolates or to their geographical origin, and no correlation between free and protein-bound BMAA concentrations were observed within or between taxonomic groups. These data offer confirmation of the taxonomic and geographic ubiquity of BMAA from naturally occurring populations of cyanobacteria, for the first time reported for estuaries.

Keywords: AQC; BMAA; Cyanobacteria; Neurotoxin; HPLC-FD; Estuaries

Importance of amino acid composition to improve skin collagen protein synthesis rates in UV-irradiated mice by Hitoshi Murakami; Kazutaka Shimbo; Yoshiko Inoue; Yoshinobu Takino; Hisamine Kobayashi (pp. 2481-2489).

Skin collagen metabolism abnormalities induced by ultraviolet (UV) radiation are the major causes of skin photoaging. It has been shown that the one-time exposure of UV irradiation decreases procollagen mRNA expression in dermis and that chronic UV irradiation decreases collagen amounts and induces wrinkle formation. Amino acids are generally known to regulate protein metabolism. Therefore, we investigated the effects of UV irradiation and various orally administered amino acids on skin collagen synthesis rates. Groups of 4–5 male, 8-week-old HR-1 hairless mice were irradiated with UVB (66 mJ/cm²) twice every other day, then fasted for 16 h. The fractional synthesis rate (FSR; %/h) of skin tropocollagen was evaluated by incorporating l-[ring-²H₅]-phenylalanine. We confirmed that the FSR of dermal tropocollagen decreased after UVB irradiation. The FSR of dermal tropocollagen was measured 30 min after a single oral administration of amino acids (1 g/kg) to groups of 5–16 UVB-irradiated mice. Branched-chain amino acids (BCAA, 1.34 ± 0.32), arginine (Arg, 1.66 ± 0.39), glutamine (Gln, 1.75 ± 0.60), and proline (Pro, 1.48 ± 0.26) did not increase the FSR of skin tropocollagen compared with distilled water, which was used as a control (1.56 ± 0.30). However, essential amino acids mixtures (BCAA + Arg + Gln, BCAA + Gln, and BCAA + Pro) significantly increased the FSR (2.07 ± 0.58, 2.04 ± 0.54, 2.01 ± 0.50 and 2.07 ± 0.59, respectively). This result suggests that combinations of BCAA and glutamine or proline are important for restoring dermal collagen protein synthesis impaired by UV irradiation.

Keywords: Amino acids; Skin collagen; Protein synthesis rate; UV-irradiated rat

Alpha-ketoglutarate inhibits glutamine degradation and enhances protein synthesis in intestinal porcine epithelial cells by Kang Yao; Yulong Yin; Xilong Li; Pengbin Xi; Junjun Wang; Jian Lei; Yongqing Hou; Guoyao Wu (pp. 2491-2500).

α-Ketoglutarate (AKG) is a key intermediate in glutamine metabolism. Emerging evidence shows beneficial effects of AKG on clinical and experimental nutrition, particularly with respect to intestinal growth and integrity. However, the underlying mechanisms are unknown. Intestinal porcine epithelial cells (IPEC-1) were used to test the hypothesis that AKG inhibits glutamine degradation and enhances protein synthesis. IPEC-1 cells were cultured for 3 days in Dulbecco’s modified Eagle’s-F12 Ham medium (DMEM-F12) containing 0, 0.2, 0.5 or 2 mM of AKG. At the end of the 3-day culture, cells were used to determine l-[U-¹⁴C]glutamine utilization, protein concentration, protein synthesis, and the total and phosphorylated levels of the mammalian target of the rapamycin (mTOR), ribosomal protein S6 kinase-1 (S6K1) and eukaryotic initiation factor (eIF) 4E-binding protein-1 (4E-BP1). Compared with 0 mM of AKG (control), 0.2 and 0.5 mM of AKG dose-dependently reduced (P < 0.05) glutamine degradation and the production of glutamate, alanine and aspartate in IPEC-1 cells. Addition of 0.5 and 2 mM of AKG to culture medium enhanced protein synthesis (P < 0.05) by 78 and 101% without affecting protein degradation, compared to the control group. Rapamycin (50 nM; a potent inhibitor of mTOR) attenuated the stimulatory effect of AKG on protein synthesis. Consistent with these metabolic data, the addition of 0.5 or 2 mM of AKG to culture medium increased (P < 0.05) the phosphorylated levels of mTOR, S6k1 and 4E-BP1 proteins. Collectively, these results indicate that AKG can spare glutamine and activate the mTOR signaling pathway to stimulate protein synthesis in intestinal epithelial cells.

Keywords: α-Ketoglutarate; Intestinal cells; mTOR signaling; Protein synthesis

Anaphylactic shock and lethal anaphylaxis caused by compound amino acid solution, a nutritional treatment widely used in China by Wen-Jian Meng; Yuan Li; Zong-Guang Zhou (pp. 2501-2505).

Compound amino acid solution (CAAS) is a large class of solution of amino acids’ mixture and was widely used in China. Its extensive nutritional treatment was accompanied by a substantial incidence of adverse reactions, especially life-threatening anaphylaxis. However, the adverse reactions were reported in isolated case reports only, and the reasons behind this needed further investigation. The Chinese language papers were searched from China National Knowledge Infrastructure and Wanfang database published in China from 1985 to 2010. The search terms “anaphylactic”, “anaphylaxis”, “allergic”, “allergy”, “shock”, and “adverse reaction” combined with the term “amino acid” were used. Totally 71 episodes of anaphylactic shock and seven deaths in 38 articles were analyzed. Chest distress and cool extremities were the most common clinical manifestations. Almost all patients suffered from significant hypotension. The vast majority of patients were not found to be allergic to certain substances. CAAS was inappropriately administrated in more than one-third of patients. The life-threatening anaphylaxis was prominently prevalent in pregnant women, the elderly and patients with hypersensitivity such as asthma, and patients without medicinal indication. Innovation of processing technique and establishment of more strict supervision system are an urgent need for CAAS to control its production quality and thus improve its safety in China.

Keywords: Anaphylactic shock; Compound amino acid solution; Lethal anaphylaxis; Nutritional treatment

The non-proteinogenic amino acids l-methionine sulfoximine and dl-phosphinothricin activate mTOR by Saverio Tardito; Martina Chiu; Renata Franchi-Gazzola; Valeria Dall’Asta; Paola Comi; Ovidio Bussolati (pp. 2507-2512).

l-Methionine sulfoximine (MSO) and dl-Phosphinothricin (PPT), two non-proteinogenic amino acids known as inhibitors of Glutamine Synthetase, cause a dose-dependent increase in the phosphorylation of the mTOR substrate S6 kinase 1. The effect is particularly evident in glutamine-depleted cells, where mTOR activity is very low, but is detectable for PPT also in the presence of glutamine. The stimulation of mTOR activity by either MSO or PPT is strongly synergized by essential amino acids. Thus, the non-proteinogenic amino acids MSO and PPT are mTOR activators.

Keywords: Glutamine; Glutamine synthetase; Methionine sulfoximine; Phosphinothricin; mTOR

Epigallocatechin-3-O-gallate inhibits the production of thymic stromal lymphopoietin by the blockade of caspase-1/NF-κB pathway in mast cells by Phil-Dong Moon; In-Hwa Choi; Hyung-Min Kim (pp. 2513-2519).

The cytokine thymic stromal lymphopoietin (TSLP) has been implicated in the development and progression of allergic diseases such as atopic dermatitis, asthma, and chronic obstructive pulmonary disease. However, it has not yet been clarified the effect of epigallocatechin-3-O-gallate (EGCG) on the production of TSLP. Thus, we investigated how EGCG inhibits the production of TSLP in the human mast cell line (HMC-1) cells. Enzyme-linked immunosorbent assay, reverse transcription-polymerase chain reaction, luciferase assay, and Western blot analysis were used to investigate the effects of EGCG. EGCG inhibited the production and mRNA expression of TSLP in HMC-1 cells. EGCG also inhibited the nuclear factor-κB luciferase activity induced by phorbol myristate acetate plus A23187. Furthermore, EGCG inhibited the activation of caspase-1 in HMC-1 cells. These results provide evidence that EGCG can help us to treat inflammatory and atopic diseases through the inhibition of TSLP.

Keywords: Thymic stromal lymphopoietin; Epigallocatechin-3-O-gallate; Nuclear factor-κB; Caspase-1

Glutamate carboxypeptidase II (GCPII) inhibitor displays anti-glutamate and anti-cocaine effects in an invertebrate assay by Chris Tallarida; Kevin Song; Robert B. Raffa; Scott M. Rawls (pp. 2521-2524).

Glutamate carboxypeptidase II (GCPII) inhibitors are promising anti-glutamatergic and anti-addictive agents. We hypothesized that a GCPII inhibitor 2 (phosphonomethyl) pentanedioic acid (2-PMPA) would display anti-stereotypical activity in planarians. Experiments revealed that 2-PMPA displayed no overt behavioral activity by itself but attenuated stereotypical counts (C-shape hyperkinesias) elicited by four compounds (2-PMPA rank order potency: glutamate > NMDA > pilocarpine > cocaine). These data suggest GCPII inhibitors display broad-spectrum efficacy against behavioral activity produced by glutamatergic and non-glutamatergic compounds in an invertebrate assay.

Keywords: GCPII; Glutamate; 2-PMPA; Cocaine; Planaria; NMDA; Pilocarpine; Stereotypy

A convenient route to optically pure α-alkyl-β-(sec-amino)alanines by A. Olma; A. Lasota; A. Kudaj (pp. 2525-2528).

The cyclization of N-Boc-α-alkylserines to corresponding β-lactones under Mitsunobu reaction conditions and the ring opening with heterocyclic amines (pyrrolidine, piperidine, morpholine and thiomorpholine) produced N-Boc-α-alkyl-β-(sec-amino)alanines. The removal of the Boc group gives di-hydrochlorides of non-protein amino acids.

Keywords: α-Alkyl-β-(sec-amino)alanines; N-Boc-α-alkylserines β-lactones; α,α-Disubstituted glycines; Mitsunobu reaction

Erratum to: Human Alzheimer’s disease synaptic O-GlcNAc site mapping and iTRAQ expression proteomics with ion trap mass spectrometry by Yuliya V. Skorobogatko; John Deuso; Jared Adolf-Bryfogle; Matthew G. Nowak; Yuesong Gong; Carol Frances Lippa; Keith Vosseller (pp. 2529-2529).

Erratum to: Preliminary kinetic characterization of a copper amine oxidase from rat liver mitochondria matrix by Roberto Stevanato; Sara Cardillo; Michele Braga; Angela De Iuliis; Valentina Battaglia; Antonio Toninello; Enzo Agostinelli; Fabio Vianello (pp. 2531-2531).

Erratum to: Chronic exposure to agmatine results in the selection of agmatine-resistant hepatoma cells by Andrea Bandino; Valentina Battaglia; Vittoria Bravoco; Chiara Busletta; Alessandra Compagnone; Carlo Cravanzola; Floriana Meli; Enzo Agostinelli; Maurizio Parola; Sebastiano Colombatto (pp. 2533-2533).

Erratum to: Polyamines modulate epithelial-to-mesenchymal transition by Alessandra Compagnone; Andrea Bandino; Floriana Meli; Vittoria Bravoco; Carlo Cravanzola; Maurizio Parola; Sebastiano Colombatto (pp. 2535-2535).

Sections

Personal tools

Amino Acids: The Forum for Amino Acid, Peptide and Protein Research (v.42, #6)

Sections

Personal tools

Local resources

Amino Acids: The Forum for Amino Acid, Peptide and Protein Research (v.42, #6)