- One-Pot Enzymatic Synthesis of d-Arylalanines Using Phenylalanine Ammonia Lyase and l-Amino Acid Deaminase
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The phenylalanine ammonia-lyase (AvPAL) from Anabaena variabilis catalyzes the amination of substituent trans-cinnamic acid (t-CA) to produce racemic d,l-enantiomer arylalanine mixture owing to its low stereoselectivity. To produce high optically pure d-arylalanine, a modified AvPAL with high d-selectivity is expected. Based on the analyses of catalytic mechanism and structure, the Asn347 residue in the active site was proposed to control stereoselectivity. Therefore, Asn347 was mutated to construct mutant AvPAL-N347A, the stereoselectivity of AvPAL-N347A for d-enantiomer arylalanine was 2.3-fold higher than that of wild-type AvPAL (WtPAL). Furthermore, the residual l-enantiomer product in reaction solution could be converted into the d-enantiomer product through stereoselective oxidation by PmLAAD and nonselective reduction by reducing agent NH3BH3. At optimal conditions, the conversion rate of t-CA and optical purity (enantiomeric excess (eeD)) of d-phenylalanine reached 82% and exceeded 99%, respectively. The two enzymes displayed activity toward a broad range of substrate and could be used to efficiently synthesize d-arylalanine with different groups on the phenyl ring. Among these d-arylalanines, the yield of m-nitro-d-phenylalanine was highest and reached 96%, and the eeD exceeded 99%. This one-pot synthesis using AvPAL and PmLAAD has prospects for industrial application.
- Zhu, Longbao,Feng, Guoqiang,Ge, Fei,Song, Ping,Wang, Taotao,Liu, Yi,Tao, Yugui,Zhou, Zhemin
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- PROTEIN MODIFIER PRODUCTION INHIBITOR
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[PLOBLEMS] To provide a inhibitor of protein modification products formation capable of inhibiting of vitamin B6 deficiency disease as a side effect, especially a renal protective agent. [MEANS FOR SOLVING PROBLEMS] There is provided a use, as an active ingredient, of any of free or salt-form compounds of either of the formulae: (I) (II) [wherein R1 is substituted or unsubstituted aromatic ring; and each of R2, R3 and R4 is a hydrogen atom or monovalent organic group, or alternatively R2 and R3 cooperate to form a condensed ring or R3 and R4 cooperate to represent a divalent organic group, provided that R3 and R4 are not simulataneously hydrogen atoms].
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Page/Page column 19; 31-32
(2008/06/13)
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- Synthesis of chirally pure 2,5-disubstituted diketopiperazines derived from trisubstituted phenylalanines
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Some new chirally pure 2,5-substituted diketopiperazines were synthesized starting from 2-methoxybenzyl alcohol. This multistep synthesis proceeds through the enzymatic synthesis of chirally pure amino acids, protection and dipeptide coupling, cyclization of dipeptide ester formates, and nitration of the resulting diketopiperazines.
- Danthi, Satyavijayan Narasimhan,Hill, Ronald A.
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p. 835 - 844
(2007/10/03)
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- Aromatic hydroxylations by flavins: Evidence on direct attack of phenylalamine by flavin radical species
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In 0.05 - 12.0 N acidic solutions, the 5-ethyl-3-methyllumiflavosemiquinone 5 (and/or 5H+) spontaneously arose from the corresponding flavinium cation 4. Raising the temperature from 20 to 50°C, considerably increased the reaction rates with no significant changes in the yields of 5 (5H+). The spontaneous one-electron reduction of 4 requires a coupling with a one-electron oxidation of another flavin such as 5-ethyl-4(a)-hydroxy-3-methyl-4(a),5-dihydroflavin pseudobase 1. The latter, being in equilibrium with 4, can be oxidized to give the transient 5-ethyl-4(a)-hydroxy-3-methyllumiflavin radical 2. This is the protonated form of a flavinoxy radical 3(a,b), a product of a homolysis of the O-O bond in a dihydroflavin hydroperoxide. As an alternative to the homolysis mentioned, the one-electron oxidation of 1 provides the principle to develop a new hydroxylating model system that does not require a dihydroflavin hydroperoxide as a starting compound. Using phenylalanine as a test substrate, the anaerobic formation of tyrosine and its o- and m-hydroxyphenylalanine isomers was established. This achievement is a strong experimental support for the hypothesis that flavin radical species like 2 may directly attack an aromatic. Evidence was obtained on some accumulation of an intermediate that is not a hydroxycyclohexadienyl radical. It was shown to react in a secondary, oxidative chain reaction, remarkably increasing the yields of aromatic hydroxylation without any further supply of flavin.
- Mager,Tu
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p. 5287 - 5298
(2007/10/02)
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- A novel hydroxylation of aromatics in a flavin-initiated chain reaction
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In aqueous acidic solutions, the 5-ethyl-3-methyllumiflavinium cation 5 was spontaneously transformed into the dihydroflavin pseudobase radical 6 and the flavosemiquinone 7. In attacking an aromatic like phenylalanine, 6 was mainly reconverted to 5 which, ultimately, could lead to an anaerobic accumulation of 7 in yields of 95 ± 5%. In the immediate presence of O2 and/or H2O2, 7 was rapidly and continuously reoxidized to 5 which, consequently, gave a continued production of 6 increasing the hydroxylating ability of the system. The oxidants also had a second distinct effect in converting an intermediate X in a chain reaction to further increase the yields of hydroxyphenylalanines. As illustrated by the results obtained with O2 and H2O2 under comparable conditions, the efficiency of this chain reaction proved to be significantly influenced by the nature of the oxidant. These new findings imply that hydroxyl radicals arising in a homolysis of the O-O bond in a dihydroflavin hydroperoxide should not be taken for granted as the primary attacking species in the hydroxylation of an aromatic. From a practical point of view it is noticed that the hydroxylating ability of the new flavin / H2O2 system surpasses that of any other known, flavin-free chemical system.
- Mager, Humphrey I. X.,Tu, Shiao-Chun
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p. 6759 - 6766
(2007/10/02)
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- HYDROXYLATION OF AROMATIC RINGS IN AN AQUEOUS SOLUTION INDUCED BY ARGON ARC PLASMA
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Plasma-induced reaction in an aqueous solution of phenyl-containing amino acids resulted in the formation of several hydroxyphenyl amino acids, which was explained by the action of hydroxyl radicals generated by decomposition of water molecules by the high energy plasma.
- Takasaki, Michiaki,Harada, Kaoru
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p. 885 - 888
(2007/10/02)
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- Studies on the Hydroxylation of Phenylalanine by 6,7-Dimethyl-5,6,7,8-tetrahydropteridine
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The hydroxylation of phenylalanine by 6,7-dimethyl-5,6,7,8-tetrahydropteridine (DMPH4) was investigated.When phenylalanine was treated with DMPH4 in citrate buffer (pH 6.0), p-tyrosine, m-tyrosine and o-tyrosine were identified as hydroxylated products.The hydroxylation was pH-dependent, and the maximum rate was found at around pH 6.Replacement of air with nitrogen gas and the addition of hydroxyl radical scavengers or catalase prevented the hydroxylation.In contrast, ferrous ions significantly accelerated the hydroxylation as compared with other transition metal ions.In an aqueous solution of DMPH4 under aerobic conditions, the electron spin resonance (ESR) spectra of the hydroxyl radical spin adducts with spin traps such as α-phenyl N-tert-butylnitrone (PBN) and α-4-pyridyl 1-oxide N-tert-butyl nitrone (4-POBN) were observed.The results indicate that the hydroxylating effect of DMPH4 is caused by hydroxyl radicals formed during the autooxidation of DMPH4.Keywords - hydroxylation; phenylalanine; p-tyrosine; m-tyrosine; o-tyrosine; 6,7-dimethyl-5,6,7,8-tetrahydropteridine; ESR; spin-trapping; hydroxyl radical
- Ishimitsu, Susumu,Fujimoto, Sadaki,Ohara, Akira
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p. 752 - 756
(2007/10/02)
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