Amino add oxidases

The natural amino acids are mainly a-amino acids, in contrast to (3-amino acids such as p-alanine and taurine. Most a-amino acids have four different substituents at C-2 (Ca). The a atom therefore represents a chiral center—I e., there are two different enantiomers (L- and D-amino acids see p. 8). Among the proteinogenic amino acids, only glycine is not chiral (R = H). In nature, it is almost exclusively L-amino acids that are found. D-Amino acids occur in bacteria—e. g., in murein (see p.40)—and in peptide antibiotics. In animal metabolism, D-Amino acids would disturb the enzymatic reactions of L-amino acids and they are therefore broken down in the liver by the enzyme D-amino add oxidase. 

B. Geueke and W. Hummel, A new bacterial L-amino add oxidase with a broad substrate spedfidty purification and charaderization, Enzyme Microb. Technol. 2002b, 31, 77-87. 

The preparation of D-amino acids with the above three-enzyme system requires enzymes with opposite stereochemical selectivity and a suitable amino add as a donor. While D-amino add oxidase is an enzyme, the function of which in Nature is mainly related to the ehmination of D-amino adds, L-amino acid oxidases are usually found in aggressive animals (snakes). Bacterial L-amino acid oxidases often show a specific activity that is too low for preparative purposes [33]. Moreover, D-amino transferases are less common than the L-specific ones and require more expensive D-amino adds as amino donors. 

Thus, using L-amino add oxidase from P. myxcfaciens and various amine-borane complexes or D-amino acid oxidase from porcine kidney and sodium cyanoboro-hydride, the preparation of several natural and non-natural enantiopure D- and L-amino adds was achieved, respectively [51]. In a more recent report, several P- and y-substituted a-amino adds were deracemized using D-amino add oxidase from Trigonopsis variahilis and sodium cyanoborohydride or sodium borohydride [52] (Scheme 13.20). 

L-Amino acid oxidase is a flavoenzyme that catalyzes the oxidative deamination of L-amino adds. L-Amino acid oxidase activities have been detected in mammals, birds, reptiles, invertebrates, molds, and bacteria [54]. L-Amino acid oxidases show the typical absorption spectrum due to the presence of a molecule of non-covalently bound FAD per subunit (with maxima at 465 and 380nm) they behave like flavoprotein oxidases, as in the case of D-amino acid oxidase. L-Amino add oxidase isolated from rat liver was reported to utilize flavin mononudeotide (FMN) as a co-enzyme, but since it is more active on L-hydroxy acids than on amino adds, it was thus considered as an L-hydroxy add oxidase. Even a partially purified L-amino acid oxidase from turkey Uver appeared to have FMN as a co-factor. 

The best substrates for ophidian L-amino acid oxidases are aromatic or, most generally, hydrophobic amino acids, polar and basic amino acids being deami-nated at much lower rates Glu, Asp, and Pro are not oxidized by L-amino add oxidase. L-Amino add oxidase is also active on ring-substituted aromatic amino acids, as well as on seleno cysteinyl derivatives. The substrate specificity depends on the source of the enzyme (e.g. Ophiophagus hannah L-amino acid oxidase also oxidizes Lys and Om) and on the pH. The of the reaction of Crotalus adaman-... 

Caligiuri, A., D Arrigo, P., Rosini, E Tessaro, D., Molla, G., Servi, S. and Polle-gioni, L. (2006) Enzymatic conversion of unnatural amino adds by yeast D-amino add oxidase. Advanced Synthesis Catalysis, 348, 2183-2190. 

L-Amino add oxidases. Small amounts of ammonia are generated by various L-amino acid oxidases, found in liver and kidney, that require a flavin mononucleotide (FMN) coenzyme. FMN is regenerated from FMNH2 by reacting with 02 to form H202. 

On the mechanism of D-amino add oxidase. Structure/linear free energy correlations and deuterium kinetic isotope effects using substituted phenylglydnes,... 

Pilone M S, Pollegioni L (2002) D-amino add oxidase as an industrial biocatalyst. Biocatal Biotransform 20 145-159... 

J. Fang, T. Sawa, T. Akaike, H. Maeda, Tumor-targeted delivery of polyethylene glycol-conjugated d-amino add Oxidase for antitumor therapy via enzymatic generation of hydrogen peroxide. Cancer Res. 62 (2002) 3138-3143. 

Snake venoms a mixture of toxins produced in the venom glands (parotid gland, or salivary gland of the upper jaw) of venomous snakes (asps or hooded snakes, e.g. the cobra sea snakes vipers, e.g. puff adder, rattlesnake). They consist of highly toxic, antigenic polypeptides and proteins (which cause paralysis and death of the prey), and enzymes (which facilitate the spread of the toxins, and initiate digestion of undivided swallowed prey). The enzymes indude hya-luronidase (promotes spread of toxins), ATPase and acetylcholine esterase (paralysis), phospholipases (hemolysis), proteinases and L-amino add oxidases (tissue necrosis and blood clotting). 

L-Amino Add Oxidase. In 1944 L-amino acid oxidases were described in 3 sources animal kidney, snake venom, and bacteria. The enzyme was also detected in animal liver. The purification from rat kidney required large amounts of tissue because of the low activity. A 200-fold purification gave a preparation believed to be essentially pure. This protein has a molecular weight somewhat over 120,000, and contains 2 flavin mononucleotides per mole. If the protein isolated is indeed the enzyme, it has one of the lowest turnover numbers determined only 6 molecules of amino acid are oxidized per molecule of protein per minute, whereas n-amino add oxidase has a turnover number of more than 1000. 

L-Amino add oxidase activity has been detected in a variety of moulds, yeasts and bacteria, but the enzymes from these sources have not been well characterised. A partly purified enzyme from Proteus vulgaris is curious in that it does not apparently produce hydrogen peroxide in the presence of molecular oxygen (equations 22-26) and absence of catalase. 

Rat Kidney -h-Amino Add Oxidase Dehydrogenase). Studies by Krebs clearly indicated that different systems existed in animal tissues for the oxidation of n- and L-amino acids. However, while considerable success has been achieved in the purification of mammahan n-amino acid... 

Mammalian-T>-Amino Add Oxidases Dehydrogenases). Since the initial demonstration by Krebs of the existence of a D-amino acid oxidase in mammalian liver and kidney, considerable progress has been made in the purification, mechanism of action, substrate specificity, etc., of this enzyme. The enzyme has been shown to be present in the liver and kidney of all vertebrates studied, but absent from other mammalian tissues. ... 

The enzymes catalysing this reaction are the L-amino add oxidases and the D-amino add oxidases. The role of the latter in metabolism has not yet been elucidated, forthenaturally occurring amino acids are generallyof the L-series. 

The optical isomers of alanine have been prepared by amination of n- and v-a-bromopropionic acids. The latter have been prepared by the action of PC1, and acetyl chlo-nde on n-serine methyl ester hydrochloride (904), by tiie action of nitroqrl bromide on n-alanine (289), and by the resolution of nir -bromopropionic add (147,6S4-656,507, 845). The preparation of n-alanine by the action of yeast on nn-alanine has been described by Ehrlich (234). L-Alanine has been prepared by the actimi of takadiastase (408, 584) and n-amino add orddase (79, 229) obtained from sheep or hog kidneys. The last author (Behrmn) isolated 22 g. of nearly pure ir-alanine from the reaction products of 72 g. of on-alanine. The velodty of the oxidation of i>-alanine by o-amino add oxidase has been determined by Stadie and Zapp (7S2). 

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