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Amino Acids: The Forum for Amino Acid, Peptide and Protein Research (v.19, #2)

Amino acid transport in the renal proximal tubule by T. Gonska; J. R. Hirsch; E. Schlatter (pp. 395-407).

In the kidney the proximal tubule is responsible for the uptake of amino acids. This occurs via a variety of functionally and structurally different amino acid transporters located in the luminal and basolateral membrane. Some of these transporters show an ion-dependence (e.g. Na⁺, Cl⁻ and K⁺) or use an H⁺-gradient to drive transport. Only a few amino acid transporters have been cloned or functionally characterized in detail so far and their structure is known, while little is known about a majority of amino acid transporters. Only few attempts have been untertaken looking at the regulation of amino acid transport. We summarized more recent information on amino acid transport in the renal proximal tubule emphasizing functional and regulatory aspects.

Keywords: Keywords: Amino acids – Kidney – IHKE-1-cells – LLC-PK1-cells – Regulation – DOG – PKC

Taurine as a universal carrier of lipid soluble vitamins: a hypothesis by A. M. Petrosian; J. E. Haroutounian (pp. 409-421).

In the literature taurine is characterized as a non-specific growth or blood clotting factor, an antioxidant, a membrane protector, or a regulator of calcium ion homeostasis, just as vitamins A, D, E, F, and K are similarly characterized. On the basis of recent finding concerning the relationship between taurine and the aldehyde of vitamin A-retinal (Petrosian and Haroutounian, 1988, 1998; Petrosian et al., 1996), as well as on the basis of data from the literature, we now suggest a hypothesis that taurine promotes the bioavailability of the lipid soluble vitamins A, D, E, K, and F, probably by forming different types of water soluble, easily hydrolyzable complexes. It is quite possible that the ability of taurine to convert lipids and lipid soluble substances into a water soluble state is the key to understanding the unusually wide diversity of biological phenomena associated with taurine. This form of delivery may be an additional, secondary mechanism for the transport of lipid soluble vitamins, which was probably acquired early in evolution, and remains extremely important for mammals and humans directly after birth for a variety of physiological functions such as: vision in normal and in emergency situations, rapid blood clotting, sperm eruption, and situations requiring a prompt consumption of lipid soluble vitamins characteristic of excitable systems. Clearly, the role of taurine in the physiology of the water insoluble vitamins remains an enigma and is worthy of further investigations.

Keywords: Keywords: Taurine – Vitamin A – Vitamin D – Vitamin E – Vitamin K – Linoleic acid

Influence of nitric oxide synthase activity on amino acid concentration in the quinolinate lesioned rat striatum: A microdialysis and histochemical study by R. Böckelmann; M. Reiser; T. F. W. Horn; G. Wolf (pp. 423-437).

The influence of nitric oxide synthase (NOS) activity on the KCl-evoked amino acid concentrations was investigated by in vivo microdialysis in the striatum in a rat model of excitotoxic lesion. Basal microdialysate levels of amino acids decreased during the quinolinic acid-induced neurodegeneration process, except for glutamine that increased initially and returned to control values 30 days after quinolinic acid exposure. KCl-evoked increase of extracellular amino acid concentration was reduced due to NOS activity in the striatum of both controls and lesioned animals, except for 120 days after quinolinic acid injection. These changes of amino acid concentrations in microdialysates correlated with the known biochemistry of the consecutive domineered cell types during the lesion process as revealed by histochemistry for NOS, NADPH-diaphorase, GFAP and isolectin B4. The present data provide direct evidence that NOS activity can modulate extracellular amino acid concentrations in the striatum not only under physiological conditions, but also during a pharmacologically induced lesion process and, thus, suggests that nitric oxide affects neurodegeneration via this pathway.

Keywords: Keywords: Amino acids – Brain microdialysis – HPLC – Immunohistochemistry – Arginine – Glutamate – Neurotoxicity

EPR study of anion radicals of various N-quinonyl amino acids by S. Bittner; S. Gorohovsky; E. Lozinsky; A. I. Shames (pp. 439-449).

Since peptide quinones possess great clinical potential in targeted chemotherapy, several series of novel N-quinonyl amino acids have been synthesized and their first products of reduction were studied by EPR spectroscopy. EPR spectra of the corresponding radical adducts were identified by computer simulation. The dependence between the splitting constants and the chemical structure of the N-quinonyl amino acids anion radicals was examined.

Keywords: Keywords: Amino acids – EPR – Semiquinone radical – N-Quinonyl amino acids – Hyperfine splittings

Relationship between NO synthesis, arginine transport, and intracellular arginine levels in vascular smooth muscle cells by N. Escobales; M. Rivera-Correa; P. I. Altieri; J. F. Rodriguez (pp. 451-468).

The present study was designed to evaluate the relevance of arginine transport in nitric oxide (NO) synthesis in vascular smooth muscle cells. For this purpose, NO synthesis and arginine transport (system B^0,+ and y⁺) were evaluated in cells treated with IL-1β or angiotensin II (Ang II). In addition, the effects of 5 mM lysine and glutamine, competitive inhibitors of systems y⁺ and B^0,+ respectively, were examined. L-arginine transport was estimated with ³H-labelled arginine and NO was determined with the Griess reagent. These studies were done in control conditions, arginine-starved cells, and in cells incubated in media containing 10 mM arginine. Our data indicate that induction of NO biosynthesis by IL-1β depends on external arginine when cells are arginine-depleted for 24 hours. The concentration of arginine producing half maximal activation of NO synthesis in arginine-depleted cells ([arginine]_i < 10 μM) was 41.1 ± 18 μM. By contrast, in normal culture conditions, NO synthesis occurred independently of arginine transport. Neither 5 mM lysine or glutamine which abolished arginine transport through systems y⁺ and B^0,+, respectively, reduced nitrite release in cells incubated in normal media. This suggests that the relevance of arginine uptake to NO synthesis depends on the status of intracellular arginine pools. Intracellular arginine concentrations were not affected by the stimulation of NO production using IL-1β or its inhibition using Ang II, but were markedly reduced by arginine starvation for 48 h. Aspartate levels were also reduced by arginine-depletion, but were not affected in cells incubated with 10 mM arginine. By contrast, glutamate levels were reduced in arginine-starved cells and were increased in cells incubated in arginine-supplemented medium. Ornithine levels were markedly increased by arginine supplementation. Altogether, these findings indicate that NO synthesis is normally independent of membrane transport. However in arginine-depleted cells, membrane transport is essential for NO synthesis. It is concluded that arginine transport is required for the long-term maintenance of intracellular arginine pools.

Keywords: Keywords: Amino acids – Nitric oxide – Angiotensin II – IL-1β– Arginine – Aspartate – Ornithine – Membrane transport – Arginine-citrulline cycle – Vascular smooth muscle cells

Diagnosis and follow-up of cystinuria: Use of proton magnetic resonance spectroscopy by G. Pontoni; F. Rotondo; G. Spagnuolo; M. T. Aurino; M. Cartenì-Farina; V. Zappia; G. Lama (pp. 469-476).

Proton Nuclear Magnetic Resonance (NMR) Spectroscopy of urine (as well as of other biological fluids) is a very powerful technique enabling multi-component analysis useful in both diagnosis and follow-up of a wide range of inherited metabolic diseases. Among these pathologies, cystinuria is characterised by accumulation in urine of four dibasic amino acids, namely lysine, arginine, ornithine and cystine; the last one, being only slightly water soluble, generates urolithiasis. The mentioned aminoacids can be detected in the urine NMR spectrum of cystinuric patients, the most abundant being the lysine (5 mM and over are often detected), whose typical signals become very high; arginine and ornithine are also usually detectable, although pathologic concentrations are lower (usually below 2 mM).The proposed NMR technique is also suitable in monitoring the therapy with α-mercaptopropionylglycine (MPG), providing quantitation of several metabolites of interest in the follow-up of the pathology, like cystine, creatinine and citrate.

Keywords: Keywords: Amino acids – Cystine – Cystinuria – Nuclear Magnetic Resonance – Urolythiasis – Urinalysis

Resolution of dl-hydantoins by d-hydantoinase from Vigna angularis: Production of highly enantioenriched N-carbamoyl-d-phenylglycine at 100% conversion by M. B. Arcuri; O. A. C. Antunes; S. J. Sabino; G. F. Pinto; E. G. Oestreicher (pp. 477-482).

d-Hydantoinase from Vigna angularis hydrolyzed rac-5-monosubstituted-hydantoins with polar and aromatic side chains and dihydrothymine but rac-5,5-disubstituted-hydantoins were not substrates of this enzyme. 5-Phenylhydantoin was the best substrate. By using this substrate, N-carbamoyl-d-phenylglycine was obtained in quantitative yield and over 98% ee.

Keywords: Keywords: Amino acids –d-Hydantoinase –N-Carbamoyl-d-phenylglycine – Substrate specificity –Vigna angularis

Inhibition of ornithine decarboxylase by α-difluoromethylornithine induces apoptosis of HC11 mouse mammary epithelial cells by T. Płoszaj; T. Motyl; W. Zimowska; J. Skierski; L. Zwierzchowski (pp. 483-496).

The effect of α-difluoromethylornithine (DFMO) on the apoptosis of HC11 mouse mammary epithelial cells was investigated. The involvement of reactive oxygen species (ROS) and Bcl-2 protein in the mechanism of apoptosis induced by ornithine decarboxylase (ODC) inhibition was also assessed. DFMO (0.1, 1 and 5 mM) induced apoptosis of HC11 cells in dose- and time-dependent manner. Apoptosis manifests itself with morphological features like: cell shrinkage, condensation of chromatin, pyknosis and fragmentation of nucleus, followed by secondary necrosis (putrosis). The decrease in the nuclear DNA contents appearing as the hypodiploidal peak sub-G₁ in the DNA histogram was not dependent on the presence of prolactin (5 μg/ml) in DFMO-treated cultures. Apoptosis induced by ODC inhibition was associated with a rapid increase in ROS concentration in HC11 cells observed within 1 h after DFMO treatment. The down-regulation of Bcl-2 as a decrease in cell number expressing bcl-2 and a lowered Bcl-2 protein content in cells expressing this protooncogene was also noted. The administration of putrescine (50 μM) lowered the number of early-apoptotic, late-apoptotic and necrotic cells. Moreover, it increased the number of cells expressing bcl-2. In conclusion, the disturbance of cellular polyamine homeostasis by inhibition of their synthesis enhances mammary epithelial cell susceptibility to apoptosis. It may occur in the mammary gland at the end of lactation, when the depletion of circulating lactogenic hormones and activation of intra-mammary apoptogenic factors expression take place.

Keywords: Keywords: Amino acids – Polyamines – Ornithine decarboxylase –α-Difluoromethylornithine – Apoptosis – Bcl-2 – Mammary epithelial cells

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Amino Acids: The Forum for Amino Acid, Peptide and Protein Research (v.19, #2)

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Amino Acids: The Forum for Amino Acid, Peptide and Protein Research (v.19, #2)