Mutation of tyrosine 332 to phenylalanine converts dopa decarboxylase into a decarboxylation-dependent oxidative deaminase

被引:62
作者
Bertoldi, M
Gonsalvi, M
Contestabile, R
Voltattorni, CB
机构
[1] Univ Verona, Fac Med & Chirurg, Sez Chim Biol, Dipartimento Sci Neurol & Vis, I-37134 Verona, Italy
[2] Univ Roma La Sapienza, Dipartimento Sci Biochim, I-00185 Rome, Italy
[3] Univ Roma La Sapienza, CNR, Ctr Mol Biol, I-00185 Rome, Italy
关键词
D O I
10.1074/jbc.M204867200
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
A flexible loop (residues 328-339), presumably covering the active site upon substrate binding, has been revealed in 3,4-dihydroxyphenylalanine decarboxylase by means of kinetic and structural studies. The function of tyrosine 332 has been investigated by substituting it with phenylalanine. Y332F displays coenzyme content and spectroscopic features identical to those of the wild type. Unlike wild type, during reactions with L-aromatic amino acids under both aerobic and anaerobic conditions, Y332F does not catalyze the formation of aromatic amines. However, analysis of the products shows that in aerobiosis, L-aromatic amino acids are converted into the corresponding aromatic aldehydes, ammonia, and CO2 with concomitant O-2 consumption. Therefore, substitution of Tyr-332 with phenylalanine results in the suppression of the original activity and in the generation of a decarboxylation-dependent oxidative deaminase activity. In anaerobiosis, Y332F catalyzes exclusively a decarboxylation-dependent transamination of L-aromatic amino acids. A role of Tyr-332 in the Calpha protonation. step that catalyzes the formation of physiological products has been proposed. Furthermore, Y332F catalyzes oxidative deamination of aromatic amines and half-transamination of D-aromatic amino acids with k(cat) values comparable with those of the wild type. However, for all the mutant-catalyzed reactions, an increase in K-m values is observed, suggesting that Y --> F replacement also affects substrate binding.
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页码:36357 / 36362
页数:6
相关论文
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