Check out our New Publishers' Select for Free Articles

Amino Acids: The Forum for Amino Acid, Peptide and Protein Research (v.37, #2)

Highlights at the gate of tryptophan catabolism: a review on the mechanisms of activation and regulation of indoleamine 2,3-dioxygenase (IDO), a novel target in cancer disease by Antonio Macchiarulo; Emidio Camaioni; Roberto Nuti; Roberto Pellicciari (pp. 219-229).

Indoleamine 2,3-dioxygenase (IDO) catalyzes the first and rate-limiting step of Kynurenine pathway along the major route of Tryptophan catabolism. The scientific interest in the enzyme has been growing since the observations of the involvement of IDO in the mechanisms of immune tolerance and in the mechanisms of tumor immuno-editing process. In view of this latter observation, in particular, preclinical studies of small molecule inhibitors of the enzyme have indicated the feasibility to thwart the immuno-editing process and to enhance the efficacy of current chemotherapeutic agents, supporting the notion that IDO is a novel target in cancer disease.This review covers the structural and conformational aspects of substrate recognition by IDO, including the catalytic mechanism and the so-far puzzling mechanisms of enzyme activation. Furthermore, we discuss the recent advances of medicinal chemistry in the field of IDO inhibitors.

Keywords: Indoleamine 2,3-dioxygenase; Immune tolerance; Cancer; Kynurenine pathway; Tryptophan metabolism

Gibberellin-regulated gene in the basal region of rice leaf sheath encodes basic helix–loop–helix transcription factor by Setsuko Komatsu; Hironori Takasaki (pp. 231-238).

Genes regulated by gibberellin (GA) during leaf sheath elongation in rice seedlings were identified using the transcriptome approach. mRNA from the basal regions of leaf sheaths treated with GA₃ was analyzed by high-coverage gene expression profiling. 33,004 peaks were detected, and 30 transcripts showed significant changes in the presence of GA₃. Among these, basic helix–loop–helix transcription factor (AK073385) was significantly upregulated. Quantitative PCR analysis confirmed that expression of AK073385 was controlled by GA₃ in a time- and dose-dependent manner. Basic helix–loop–helix transcription factor (AK073385) is therefore involved in the regulation of gene expression by GA₃.

Keywords: Gibberellin; High-coverage gene expression profiling; Rice; Basal region of leaf sheath; Basic helix–loop–helix transcription factor

Mechanism of the suppression against d-galactosamine-induced hepatic injury by dietary amino acids in rats by Torai Komano; Reiko (Yokoyama) Funakoshi; Yukari Egashira; Hiroo Sanada (pp. 239-247).

To elucidate the mechanism by which dietary amino acids suppress the d-galactosamine (d-GalN)-induced hepatitis, we examined the involvement of Kupffer cells, tumor necrosis factor-α (TNF-α) and apoptosis in the mechanism. In experiment 1, the rats were fed with 10%l-glutamine or 5% glycine diet injected with d-GalN with or without gadolinium chloride (GdCl₃)-pretreatment. The results indicated that these amino acids suppressed the d-GalN-induced elevation of serum transaminase activities, irrespective of GdCl₃-pretreatment. In experiment 2, rats were fed with 10% of l-glutamine, l-serine, l-alanine or l-glutamic acid diets injected with d-GalN. The results demonstrated that all these amino acids suppressed the d-GalN-induced elevation of serum transaminase activities, but that serum TNF-α concentrations and hepatic caspase-3 activities in the rats were not appreciably changed. In conclusion, the suppressive effects of amino acids on d-GalN-induced hepatitis were suggested not to be always mediated by the inhibition of Kupffer cells → TNF-α → apoptosis pathway.

Keywords: l-Alanine; Gadolinium chloride; d-Galactosamine; l-Glutamic acid; l-Glutamine; l-Serine

Cellular uptake of cationic gadolinium-DOTA peptide conjugates with and without N-terminal myristoylation by A. Sturzu; U. Klose; H. Echner; A. Beck; A. Gharabaghi; H. Kalbacher; S. Heckl (pp. 249-255).

Cellular and nuclear uptake of dual labelled conjugates could be of great value for chemotherapy and cancer diagnostics. Therefore we designed conjugates in which gadolinium (Gd)-1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA), a contrast agent for magnetic resonance imaging and fluorescein isothiocyanate (FITC), a fluorescence marker were coupled to membrane translocation sequences (MTS). The MTSs we employed were the third helix of the Antennapedia homeodomain, the HIV-1 Tat peptide and the N-myristoylated HIV-1 Tat peptide. We used confocal laser scanning microscopy, fluorescence activated cell sorting, magnetic resonance imaging (MRI) and viability tests to examine the cellular and nuclear uptake of these conjugates into U373 glioma cells, as well as their cytotoxic effects. We found that the Antennapedia conjugate was taken up by no more than 20% of the cells. The HIV-1 Tat conjugate showed even lower uptake into less than 3% of cells. Interestingly, N-myristoylation of the HIV-1 Tat conjugate drastically improved its cellular uptake. Up to 70% of cells showed cellular and nuclear uptake of the N-myristoylated HIV-1 Tat conjugate. Conjugate cytotoxicity appears to correlate with cellular uptake.

Keywords: N-myristoylation; HIV-1 Tat; Penetratin; Tetraazacyclododecane-tetraacetic acid; Gadolinium; Cellular uptake; Chemotherapy

Skp2 inversely correlates with p27 and tuberin in transformed cells by M. Rosner; M. Hanneder; N. Siegel; A. Valli; C. Fuchs; M. Hengstschläger (pp. 257-262).

The cyclin-dependent kinase inhibitor p27^Kip1 (p27) is a major gatekeeper of the mammalian cell cycle progression known to be regulated by both, its subcellular localization and its degradation. To allow entrance into S phase and thereby mammalian cell cycle progression p27 must be degraded by a skp2-containing E3 ubiquitin ligase whose task is to target p27 for degradation by the proteasome. The tumor suppressor gene product tuberin directly binds to p27 and protects it from degradation via skp2. Whereas, p27 and tuberin are known to be localized to both, the cytoplasm and the nucleus, the localization of skp2 remained elusive. Here we demonstrate that skp2 is a cytoplasmic and nuclear protein. In addition we found an inverse correlation of the endogenous protein levels of skp2 with p27 and tuberin in different transformed cells and under different growth conditions. These data allow new important insights into this molecular network of cell cycle control.

Keywords: Skp2; Tuberin; p27

Dietary supplementation with Astragalus polysaccharide enhances ileal digestibilities and serum concentrations of amino acids in early weaned piglets by F. G. Yin; Y. L. Liu; Y. L. Yin; X. F. Kong; R. L. Huang; T. J. Li; G. Y. Wu; Yongqing Hou (pp. 263-270).

Two experiments were conducted to evaluate the effects of dietary supplementation with Astragalus polysaccharide (APS) on growth performance, apparent ileal digestibilities (AID) of amino acids (AA), and their serum concentrations in early weaned piglets. In Exp. 1, 60 pigs were weaned at 21 days of age (BW 7.35 ± 0.23 kg) and allocated to three treatments (20 pigs/treatment), representing supplementing 0.0% (control), 0.02% colistin (antibiotic), or 0.1% APS to a corn- and soybean meal-based diet. Average daily gain (ADG), average daily feed intake (ADFI), and feed/gain ratio (F/G) were measured weekly. Blood samples were obtained from five pigs selected randomly from each treatment for the measurement of serum free AA concentrations on days 7, 14, and 28. In Exp. 2, 12 pigs were weaned at 21 day of age (BW 7.64 ± 0.71 kg), assigned to three treatment groups as in Exp. 1, and surgically fitted with a simple T-cannula at the terminal ileum. Ileal digesta samples were obtained for the measurement of AID of AA on days 7, 14 and 28. Dietary APS did not affect ADFI, but enhanced (P < 0.05) ADG by 11 and 4.4%, and improved F/G by 5.6 and 8.4%, respectively, compared with the control and antibiotic groups. Addition of APS to the diet increased AID and serum concentrations of most nutritionally essential and non-essential AA (including arginine, proline, glutamate, lysine, methionine, tryptophan, and threonine) on days 14 and 28. Circulating levels of total AA were affected by the age of pigs and treatment × time interaction. Collectively, these findings indicate that APS may ameliorate the digestive and absorptive function and regulate AA metabolism to beneficially increase the entry of dietary AA into the systemic circulation, which provide a mechanism to explain the growth-promoting effect of APS in early weaned piglets.

Keywords: Amino acids; Astragalus polysaccharide; Digestibilities; Performance; Pig

Glutamate is the chemotaxis-inducing factor in placental extracts by Rahul Gupta; Dhrubajyoti Chattopadhyay (pp. 271-277).

Human placental extracts are known to help wound healing. Rapid migration of neutrophils to the wound site is a prerequisite to the wound healing process. Gel filtration analysis of heat-treated placental extract gave the initial cue to the small nature of the migration promoting factor of the extract. HPLC analysis of the extract revealed glutamate to be the predominant free amino acid. Our studies show that glutamate at an optimum concentration of 8 μM induced phenotypic neutrophil chemotaxis, as seen in the time lapse- and transwell assays. Glutamate was also found to induce chemokinesis of the neutrophil, though the stimulation of chemotaxis was more pronounced. The glutamate induced chemotaxis was accompanied by polarization of the actin cytoskeleton, and by polymerization of F-actin. These data indicate that glutamate has a strong chemotactic functionality in the neutrophil, which could be of interest both therapeutically and in further investigation of the molecular basis of chemotaxis.

Keywords: Chemoattractant; Migration; Polarization; Neurotransmitter

Glutamate is the chemotaxis-inducing factor in placental extracts by Rahul Gupta; Dhrubajyoti Chattopadhyay (pp. 271-277).

Keywords: Chemoattractant; Migration; Polarization; Neurotransmitter

Variation in basal heat shock protein 70 is correlated to core temperature in human subjects by Marie E. Sandström; Leigh A. Madden; Lee Taylor; Jason C. Siegler; Ric J. Lovell; Adrian Midgley; Lars McNaughton (pp. 279-284).

Heat shock proteins are highly conserved proteins and play an important chaperone role in aiding the folding of nascent proteins within cells. The heat shock protein response to various stressors, both in vitro and in vivo, is well characterised. However, basal levels of heat shock protein 70 (Hsp70) have not previously been investigated. Monocyte-expressed Hsp70 was determined every 4 h, over a 24 h time period, in 17 healthy male subjects (177 ± 6.4 cm, 75.7 ± 10.9 kg, 19.8 ± 4.3 years) within a temperature and activity controlled environment. Core temperature was measured at 5-min intervals during the 24 h period. Hsp70 showed significant diurnal variation (F = 7.4; p < 0.001), demonstrating peaks at 0900 and 2100 hours, and a nadir at 05.00. Core temperature followed a similar temporal trend (range = 35.96–38.10°C) and was significantly correlated with Hsp70 expression (r _s = 0.44; p < 0.001). These findings suggest a high responsiveness of Hsp70 expression in monocytes to slight variations in core temperature.

Keywords: Hsp70; Diurnal variation; Core temperature

Synthesis and bioactivity of chemotactic tetrapeptides: fMLF-OMe analogues incorporating spacer aminoacids at the lateral positions by Gino Lucente; Cesare Giordano; Anna Sansone; Domenica Torino; Susanna Spisani (pp. 285-295).

A small library of N-For and N-Boc tetrapeptidic analogues of the chemotactic tripeptide For-Met-Leu-Phe-OMe (fMLF-OMe), obtained by incorporating three different spacer aminoacids (Gly, βAla and Pro) between the native residues of Met and Leu (N-For- and N-Boc-Met-Xaa-Leu-Phe-OMe; Xaa² series) and Leu and Phe (N-For- and N-Boc-Met-Leu-Xaa-Phe-OMe; Xaa³ series), have been synthesized and examined for their biological activity as agonists and antagonists. Chemotaxis, lysozyme release and superoxide anion production have been measured. All the N-For analogues maintain good to moderate chemotactic activity with the βAla³ 15 model reaching the maximum value. All the N-Boc tetrapeptides are efficient chemotactic antagonists. Conversely, with the exception of the moderate antagonistic activity exhibited by the N-Boc Xaa² models against lysozyme release, all the other N-Boc analogues do not show significant activity against both superoxide anion and lysozyme release.

Keywords: Chemotaxis; fMLF analogues; Formylpeptides; Human neutrophils; Tetrapeptides

Acute and long-term effects of resistance exercise with or without protein ingestion on muscle hypertrophy and gene expression by Juha J. Hulmi; Vuokko Kovanen; Harri Selänne; William J. Kraemer; Keijo Häkkinen; Antti A. Mero (pp. 297-308).

The effects of timed ingestion of high-quality protein before and after resistance exercise are not well known. In this study, young men were randomized to protein (n = 11), placebo (n = 10) and control (n = 10) groups. Muscle cross-sectional area by MRI and muscle forces were analyzed before and after 21 weeks of either heavy resistance training (RT) or control period. Muscle biopsies were taken before, and 1 and 48 h after 5 × 10 repetition leg press exercise (RE) as well as 21 weeks after RT. Protein (15 g of whey both before and after exercise) or non-energetic placebo were provided to subjects in the context of both single RE bout (acute responses) as well as each RE workout twice a week throughout the 21-week-RT. Protein intake increased (P ≤ 0.05) RT-induced muscle cross-sectional area enlargement and cell-cycle kinase cdk2 mRNA expression in the vastus lateralis muscle suggesting higher proliferating cell activation response with protein supplementation. Moreover, protein intake seemed to prevent 1 h post-RE decrease in myostatin and myogenin mRNA expression but did not affect activin receptor IIb, p21, FLRG, MAFbx or MyoD expression. In conclusion, protein intake close to resistance exercise workout may alter mRNA expression in a manner advantageous for muscle hypertrophy.

Keywords: cdk2; Myostatin; Activin receptor IIb; Skeletal muscle; Whey

Synthesis of gem-disubstituted taurines by the regioselective ring-opening of 2,2-disubstituted aziridines with sodium bisulfite and sulfite by Ning Chen; Min Zhu; Wei Zhang; Da-Ming Du; Jiaxi Xu (pp. 309-313).

The regioselectivity of the ring-opening reactions of 2,2-disubstituted aziridines with sodium bisulfite and sulfite showed significant divergence depending on the nucleophiles and the structure of the substituents of the aziridines. 2,2-Dialkylaziridines were specifically attacked on their less substituted carbon atoms with sodium bisulfite, affording 2,2-substituted taurines, while 2,2-disubstituted aziridines with either one or two aryl substituent(s) were attacked specifically on their more substituted carbon atoms with the same nucleophile, giving rise to aromatic 1,1-substituted taurines. Sodium sulfite attacked the less substituted carbon atoms of all aziridines specifically to afford 2,2-disubstituted taurines. The regioselectivity is governed by the nucleophile and the balance between the steric hindrance and the electronic effect. The current method provides an alternative route to the synthesis of 2,2-dialkyltaurines and aromatic gem-disubstituted taurines.

Keywords: Amino acid; Aminoalkanesulfonic acid; Ring-opening reaction; Sodium bisulfite; Sodium sulfute; Synthesis; Taurine

Laccase-catalyzed carbon–nitrogen bond formation: coupling and derivatization of unprotected l-phenylalanine with different para-hydroquinones by V. Hahn; A. Mikolasch; K. Manda; D. Gördes; K. Thurow; F. Schauer (pp. 315-321).

Unprotected l-phenylalanine was derivatized by an innovative enzymatic method by means of laccases from Pycnoporus cinnabarinus and Myceliophthora thermophila. During the incubation of l-phenylalanine with para-hydroquinones using laccase as biocatalyst, one or two main products were formed. Dependent on the substitution grade of the hydroquinones mono- and diaminated products were detected. Differences of the used laccases are discussed. The described reactions are of interest for the derivatization of amino acids and a synthesis of pharmacological-active amino acid structures in the field of white biotechnology.

Keywords: Laccase; Cross-coupling; Phenylalanine; Derivatization; Unprotected amino acid; Quinone

Dietary l-arginine supplementation enhances the immune status in early-weaned piglets by Bie Tan; Xinguo G. Li; Xiangfeng Kong; Ruilin Huang; Zheng Ruan; Kang Yao; Zeyuan Deng; Mingyong Xie; Izuru Shinzato; Yulong Yin; Guoyao Wu (pp. 323-331).

This study was conducted to test the hypothesis that dietary L-arginine supplementation enhances immunity in early weaned piglets. Seventy piglets weaned at 7 days of age were assigned to five groups (14 pigs/group), representing supplementation of 0.0, 0.2, 0.4, 0.6, and 0.8% l-arginine to a milk-based formula. On Day 7 after initiation of treatment, spleen weight in piglets supplemented with 0.2 and 0.8% arginine was heavier and thymus size was larger in piglets supplemented with 0.6% arginine, whereas serum concentration of immunoglobulin (Ig) M was higher but that of IL-8 was lower in piglets supplemented with 0.6 and 0.8% arginine, compared with the control group. Dietary supplementation with 0.8% arginine increased the numbers of white blood cells and granulocytes, and gene expression of interleukin (IL)-8 in spleen. On Day 14, compared with control piglets, granulocyte numbers were greater but lymphocyte numbers were lower in piglets supplemented with 0.2 and 0.4% arginine, whereas splenic expression of IL-8 and tumor necrosis factor-α genes was increased in piglets supplemented with 0.8% arginine. Additionally, IgG and IgM concentrations in serum and growth performance were greater in piglets supplemented with 0.4–0.8% arginine, compared with unsupplemented piglets. Collectively, dietary supplementation with 0.4–0.8% l-arginine for 2 weeks enhances both cellular and humoral immunity in piglets by modulating the production of leukocytes, cytokines and antibodies. These results indicate that increasing l-arginine provision is beneficial for optimal immune responses in young pigs and also have important implications for designing the next generation of improved formula for human infants.

Keywords: Arginine; Immunity; Metabolism; Piglet

Human insulinotropic response to oral ingestion of native and hydrolysed whey protein by O. Power; A. Hallihan; P. Jakeman (pp. 333-339).

The insulinotropic response to the ingestion of whey protein and whey protein hydrolysate, independent of carbohydrate, is not known. This study examined the effect of protein hydrolysis on the insulinotropic response to the ingestion of whey protein. Sixteen healthy males ingested a 500 mL solution containing either 45 g of whey protein (WPI) or whey protein hydrolysate (WPH). The estimated rate of gastric empting was not altered by hydrolysis of the protein [18 (3) vs. 23 (3) min, n = 16; P = 0.15]. Maximum plasma insulin concentration (C _max) occurred later (40 vs. 60 min) and was 28% [234 (26) vs. 299 (31) mM, P = 0.018] greater following ingestion of the WPH compared to the WPI leading to a 43% increase [7.6 (0.9) vs. 10.8 (2.6) nM, P = 0.21] in the AUC of insulin for the WPH. Of the amino acids with known insulinotropic properties only Phe demonstrated a significantly greater maximal concentration [C _max; 65 (2) vs. 72 (3) μM, n = 16; P = 0.01] and increase (+22%) in AUC following ingestion of the WPH. In conclusion, ingestion of whey protein is an effective insulin secretagogue. Hydrolysis of whey protein prior to ingestion augments the maximal insulin concentration by a mechanism that is unrelated to gastric emptying of the peptide solution.

Keywords: Amino acids; Insulin; Whey protein; Peptide hydrolysate; Gastric emptying

Derivatized graphitic nanofibres (GNF) as a new support material for mass spectrometric analysis of peptides and proteins by Andreas Greiderer; Matthias Rainer; Muhammad Najam-ul-Haq; Rainer M. Vallant; Christian W. Huck; Günther K. Bonn (pp. 341-348).

Graphitic nanofibres (GNFs), 100–200 nm in diameter and 5–20 μm in length have been modified in order to yield different affinities (Cu²⁺ and Fe³⁺ loaded immobilized metal affinity chromatography (IMAC) as well as cation and anion exchange materials) for the extraction of a range of biomolecules by their inherited hydrophobicity and the hydrophilic chemical functionalities, obtained by derivatization. Modified GNFs have for the first time been employed as carrier materials for protein profiling in material-enhanced laser desorption/ionization (MELDI) for the enrichment and screening of biofluids. For that purpose, the derivatized GNF materials have comprehensively been characterized regarding surface area, structural changes during derivatization, IMAC, as well as ion exchange and protein-loading capacity and recovery. GNF derivatives revealed high protein-binding capacity (2,000 μg ml⁻¹ for insulin) and ideal sensitivities, resulting in a detection limit of 50 fmol μl⁻¹ (for insulin), which is crucial for the detection of low abundant species in biological samples. Compared to other MELDI carrier materials, sensitivity was enhanced on GNF derivatives, which might be ascribed to the fact that GNFs support desorption and ionization mechanisms and by absorbing laser energy in addition to matrix.

Keywords: Nanomaterials; Graphitic nanofibres (GNF); Derivatization; Mass spectrometry (MS); Matrix-assisted laser desorption/ionization (MALDI); Material-enhanced laser desorption/ionization (MELDI); Protein profiling

Pharmacokinetics of acute tryptophan depletion using a gelatin-based protein in male and female Wistar rats by L. A. W. Jans; C. K. J. Lieben; L. T. Smits; A. Blokland (pp. 349-357).

The essential amino acid tryptophan is the precursor of the neurotransmitter serotonin. By depleting the body of tryptophan, brain tryptophan and serotonin levels are temporarily reduced. In this paper, several experiments are described in which dose and treatment effects of acute tryptophan depletion (ATD) using a gelatin-based protein–carbohydrate mixture were studied in male and female Wistar rats. Two or three doses of tryptophan depleting mixture resulted in 65–70% depletion after 2–4 h in males. ATD effects were similar in females, although females may return to baseline levels faster. Treatment effects after four consecutive days of ATD were similar to the effects of 1 day of treatment. Object recognition memory was impaired 2, 4, and 6 h after the first of two doses of ATD, suggesting that the central effects occurred rapidly and continued at least 6 h, in spite of decreasing treatment effects on plasma tryptophan levels at that time point. The method of acute tryptophan depletion described here can be used to study the relationship between serotonin and behaviour in both male and female rats.

Keywords: Serotonin; Tryptophan; Sex; Amino acids; Memory; Rat

Glutamate is the chemotaxis-inducing factor in placental extracts by Rahul Gupta; Dhrubajyoti Chattopadhyay (pp. 359-366).

Human placental extracts are known to help wound healing. Rapid migration of neutrophils to the wound site is a prerequisite to the wound healing process. Gel filtration analysis of heat-treated placental extract gave the initial cue to the small nature of the migration promoting factor of the extract. HPLC analysis of the extract revealed glutamate to be the predominant free amino acid. Our studies show that glutamate at an optimum concentration of 8 μM induced phenotypic neutrophil chemotaxis, as seen in the time lapse and transwell assays. Glutamate was also found to induce chemokinesis of the neutrophil, though the stimulation of chemotaxis was more pronounced. The glutamate induced chemotaxis was accompanied by polarization of the actin cytoskeleton, and by polymerization of F-actin. These data indicate that glutamate has a strong chemotactic functionality in the neutrophil, which could be of interest both therapeutically and in further investigation of the molecular basis of chemotaxis.

Keywords: Chemoattractant; Migration; Polarization; Neurotransmitter

Glutamate is the chemotaxis-inducing factor in placental extracts by Rahul Gupta; Dhrubajyoti Chattopadhyay (pp. 359-366).

Human placental extracts are known to help wound healing. Rapid migration of neutrophils to the wound site is a prerequisite to the wound healing process. Gel filtration analysis of heat-treated placental extract gave the initial cue to the small nature of the migration promoting factor of the extract. HPLC analysis of the extract revealed glutamate to be the predominant free amino acid. Our studies show that glutamate at an optimum concentration of 8 μM induced phenotypic neutrophil chemotaxis, as seen in the time lapse and transwell assays. Glutamate was also found to induce chemokinesis of the neutrophil, though the stimulation of chemotaxis was more pronounced. The glutamate induced chemotaxis was accompanied by polarization of the actin cytoskeleton, and by polymerization of F-actin. These data indicate that glutamate has a strong chemotactic functionality in the neutrophil, which could be of interest both therapeutically and in further investigation of the molecular basis of chemotaxis.

Keywords: Chemoattractant; Migration; Polarization; Neurotransmitter

Mutant rat strain lacking d-amino-acid oxidase by Ryuichi Konno; Tadashi Okamura; Noriyuki Kasai; Karl H. Summer; Akira Niwa (pp. 367-375).

d-Amino-acid oxidase (DAO) is known to be associated with schizophrenia. Since the expression of DAO gene had been reported to be very low in LEA rats, we examined LEA/SENDAI rats in detail. These rats did not have DAO activity, enzyme protein or mRNA encoding this enzyme. Sequencing of the 5′-upstream region of the DAO gene revealed the deletion of one triplet in the 15 TAA repeats approximately 700-bp upstream of the transcription start point. A 1.3-kb upstream fragment containing the TAA repeats and the transcription start point was inserted into a reporter vector and was transfected into COS-1, NRK-52E and CCL-PK1 cells. Although the fragments containing 15 or 14 repeats had high promoter activity, the fragment containing 13 repeats had very weak activity. Electrophoretic mobility-shift assays showed that the nuclear extracts from COS-1 and COS-7 cells had proteins that bound to the oligonucleotides containing the TAA repeats. These results suggest that the TAA repeats are important for expression of the DAO gene. The LEA/SENDAI rats lacking DAO would be a useful tool for the investigations aimed at the elucidation of the relationships between this flavoenzyme and schizophrenia.

Keywords: d-Amino-acid oxidase; Rat mutant; Promoter; Triplet repeats; Schizophrenia

l-Proline is a sedative regulator of acute stress in the brain of neonatal chicks by K. Hamasu; T. Haraguchi; Y. Kabuki; N. Adachi; S. Tomonaga; H. Sato; D. M. Denbow; M. Furuse (pp. 377-382).

The purpose of the present study was to clarify the central nervous system function of amino acids during acute stress. In Experiment 1, changes in free amino acid pattern were investigated in the brain of neonatal chicks exposed to either restraint with isolation-induced or fasting stress. l-Proline and l-arginine were decreased in the telencephalon and diencephalon under any stress. Since the central nervous system functions of l-arginine during the stress response has recently been reported, in Experiment 2, the effect of intracerebroventricular injection of l-proline (0.5, 1.0, 2.0 μmol) during isolation-induced stress was investigated. l-Proline induced sedative and hypnotic effects in a dose-dependent manner. It is suggested that l-proline may have an important role to attenuate the stress response in the central nervous system of chicks.

Keywords: l-Proline; Amino acid; Chick; Stress; Brain; i.c.v. injection

Synthesis, antioxidative and antiviral activity of hydroxycinnamic acid amides of thiazole containing amino acid by Ivanka Stankova; Kiril Chuchkov; Stoyan Shishkov; Kalina Kostova; Luchia Mukova; Angel S. Galabov (pp. 383-388).

The synthesis and the biological (antioxidant and antiviral) activities of novel hydroxycinnamic acid amides of a thiazole containing TFA.valine-4-carboxylic acid ethyl ester are reported. The amides have been synthesized from p-coumaric, ferulic and sinapic acids with the corresponding TFA.valine-thiazole-4-carboxylic acid ethyl ester using the coupling reagent N-ethyl-N′-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) and 4-(dimethylamino) pyridine (DMAP) as a catalyst. The antioxidant properties of the newly synthesized amides have been studied for then antioxidative activity using 2,2-diphenyl-1-picrylhydrazyl (DPPH)* test. The newly synthesized compounds have been tested against the replication in vitro of influenza virus A (H3N2) and human herpes virus 1 and 2 (HSV-1 and HSV-2).

Keywords: Hydroxycinnamides; Thiazole containing valine; Peptidomimetics; Antioxidant effect; DPPH test; Antiviral activity

Novel mastoparan and protonectin analogs isolated from a solitary wasp, Orancistrocerus drewseni drewseni by Kazuya Murata; Tetsuro Shinada; Yasufumi Ohfune; Miki Hisada; Akikazu Yasuda; Hideo Naoki; Terumi Nakajima (pp. 389-394).

Two novel biologically active peptides, Eumenine mastoparan-OD and Orancis-Protonectin, were isolated from a solitary wasp, Orancistrocerus drewseni drewseni (Eumeninae, Vespidae). MALDI-TOF MS analysis of a small amount of the crude venom gave two intensive molecular-related ion peaks at m/z 1269.9 and 1552.9 that were expected to be novel based on a peptide database search. Purification of the crude venom by HPLC gave two peptide fractions, P-1 and P-2. The amino acid sequence of P-1 (GRILSFIKAGLAEHL-NH₂) and P-2 (ILGIITSLLKSL-NH₂) were determined by ESI-MS/MS, automated Edman degradation, and amino acid analysis. According to the high sequence homology with those of mastoparan and protonectin, P-1 and P-2 were labeled Eumenine mastoparan-OD and Orancis-Protonectin, respectively. Orancis-Protonectin is the first example of a protonectin analog isolated from the venom of a solitary wasp. The hemolytic activities of these new peptides were more potent than that of mastoparan.

Keywords: Solitary wasp; Mass spectrometry; Mastoparan; Protonectin; Hemolytic activity

Simultaneous detection of N-acetyl-l-cysteine and physiological low molecular mass thiols in plasma by capillary electrophoresis by Angelo Zinellu; Salvatore Sotgia; Bastianina Scanu; Maria Franca Usai; Alessandro Giuseppe Fois; Valentina Spada; Anna Deledda; Luca Deiana; Piero Pirina; Ciriaco Carru (pp. 395-400).

N-acetyl-l-cysteine (NAC) is a therapeutic drug widely used as mucolytic agent in the treatment of respiratory diseases. Recently it has been proposed that NAC administration may modify the plasma levels of low molecular weight thiols (LMW) like cysteine, homocysteine and glutathione, though it has been still debated if their plasma concentration increases or decreases during the therapy. Therefore research calls for methods able to analyze simultaneously NAC and the other plasma LMW thiols in order to evaluate if NAC is able to modify plasma thiols concentration and in particular to reduce homocysteine levels in hyperhomocysteinemia. In this paper we present a new capillary electrophoresis method that allows a baseline separation of plasma NAC from the physiological thiols. The proposed method has been utilized to measure the drug and the physiological LMW thiols in NAC administered chronic obstructive broncho-pneumopathy (COPB) disease patients.

Keywords: NAC; LMW thiols; Capillary electrophoresis

A conceptual model of the polyamine binding site of N ¹-acetylpolyamine oxidase developed from a study of polyamine derivatives by Koichi Takao; Satoko Shibata; Tomohiro Ozawa; Makiko Wada; Yoshiaki Sugitia; Keijiro Samejima; Akira Shirahata (pp. 401-405).

We used various polyamine derivatives to study the substrate binding site of N ¹-acetylpolyamine oxidase (PAO) that was partially purified from rat liver. The substrate activities of acetylpolyamines indicated the presence of two anionic centers corresponding to the 1,3-diaminopropane (1,3-DAP) structure and a hydrophobic region in addition to the cleavage site of the acetamidopropyl group. Based on the results of the inhibitory activities of 1,3-DAP derivatives, we developed a conceptual model of the polyamine binding site of PAO. We used this model to identify a potent competitive inhibitor, N ¹,N ⁷-dihexyl-1,7-diamino-4-azaheptane, and to develop an affinity column, 1,16-diamino4,13-diazahexadecane–linked Sepharose, which was useful for the purification of PAO.

Keywords: Polyamines; Acetylpolyamine oxidase; Diacetylpolyamines; Acetylpolyamines; Affinity column

Altered urinary polyamine patterns of cancer patients under acupuncture therapy by M. J. Paik; D. Kuon; J. Cho; K.-R. Kim (pp. 407-413).

The reduction of elevated polyamine (PA) levels in biological fluids of cancer patients were known to be correlated with remission following diverse therapeutic treatments. In this study, altered urinary PA levels from three different cancer cases were monitored at different intervals during the long-term weekday acupuncture treatments. Nine urinary PA levels from 16 normal and three cancer patients with different types were measured by gas chromatography–mass spectrometry in selected ion monitoring mode as N-ethoxycarbonyl-N-pentafluoropropionyl derivatives. Their levels measured at three follow-up stages for each patient were then normalized to the corresponding normal group means and plotted into star symbol patterns. Large alterations of PA levels were observed for each patient. Each normalized concentration displayed elevation of the PA levels in multiples (0.0–57.7) of the respective normal mean values. The normalized PA values were transformed into distorted star patterns which were characteristic of each follow-up stage and of cancer type.

Keywords: Cancer; Acupuncture; Urinary polyamines; Ethoxycarbonyl-pentafluoropropionyl derivatives; Gas chromatography–mass spectrometry in selected ion monitoring; Star pattern recognition

Prediction of protein structural classes by Chou’s pseudo amino acid composition: approached using continuous wavelet transform and principal component analysis by Zhan-Chao Li; Xi-Bin Zhou; Zong Dai; Xiao-Yong Zou (pp. 415-425).

A prior knowledge of protein structural classes can provide useful information about its overall structure, so it is very important for quick and accurate determination of protein structural class with computation method in protein science. One of the key for computation method is accurate protein sample representation. Here, based on the concept of Chou’s pseudo-amino acid composition (AAC, Chou, Proteins: structure, function, and genetics, 43:246–255, 2001), a novel method of feature extraction that combined continuous wavelet transform (CWT) with principal component analysis (PCA) was introduced for the prediction of protein structural classes. Firstly, the digital signal was obtained by mapping each amino acid according to various physicochemical properties. Secondly, CWT was utilized to extract new feature vector based on wavelet power spectrum (WPS), which contains more abundant information of sequence order in frequency domain and time domain, and PCA was then used to reorganize the feature vector to decrease information redundancy and computational complexity. Finally, a pseudo-amino acid composition feature vector was further formed to represent primary sequence by coupling AAC vector with a set of new feature vector of WPS in an orthogonal space by PCA. As a showcase, the rigorous jackknife cross-validation test was performed on the working datasets. The results indicated that prediction quality has been improved, and the current approach of protein representation may serve as a useful complementary vehicle in classifying other attributes of proteins, such as enzyme family class, subcellular localization, membrane protein types and protein secondary structure, etc.

Keywords: Pseudo-amino acid composition; Support vector machine; Wavelet power spectrum

A complexity-based method for predicting protein subcellular location by Xiaoqi Zheng; Taigang Liu; Jun Wang (pp. 427-433).

A complexity-based approach is proposed to predict subcellular location of proteins. Instead of extracting features from protein sequences as done previously, our approach is based on a complexity decomposition of symbol sequences. In the first step, distance between each pair of protein sequences is evaluated by the conditional complexity of one sequence given the other. Subcellular location of a protein is then determined using the k-nearest neighbor algorithm. Using three widely used data sets created by Reinhardt and Hubbard, Park and Kanehisa, and Gardy et al., our approach shows an improvement in prediction accuracy over those based on the amino acid composition and Markov model of protein sequences.

Keywords: Protein subcellular location; Symbol sequence complexity; k-Nearest neighbor algorithm; Jackknife analysis

Pseudodeficiency of glutamine in infant liver disease by T. Vermeulen; T. Marquardt; J. Häberle (pp. 435-437).

γ-Glutamyltransferase (γ-GT) is an early marker for cholestasis and has the capability of glutamine-deamidation. Two infants with elevated serum γ-GT had a decreased serum glutamine. A time course of glutamine and glutamate concentration changes was performed. This revealed a time dependent decrease of glutamine far below the normal lower limit while glutamate increased above the normal upper limit. In conclusion, increased in vitro γ-GT can cause pseudodeficiency of glutamine. To avoid pitfalls, physicians should inform the laboratory on accompanying pathologies.

Keywords: γ-Glutamyltransferase; γ-GT; Glutamine deficiency; Cholestasis

Eye lens proteomics by W. Hoehenwarter; J. Klose; Peter R. Jungblut (pp. 439-439).

Sections

Personal tools

Amino Acids: The Forum for Amino Acid, Peptide and Protein Research (v.37, #2)

Sections

Personal tools

Local resources

Amino Acids: The Forum for Amino Acid, Peptide and Protein Research (v.37, #2)