Inactivation of microbial arginine deiminases by L-canavanine
Inactivation of microbial arginine deiminases by L-canavanine
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DOI：
10.1021/ja0760877
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发表时间：
2008-02-13
影响因子：
15
通讯作者：
Zhang, Liwen
中科院分区：
文献类型：
--
作者：
Li, Ling;Li, Zhimin;Zhang, Liwen
精氨酸脱节酶（ADI）催化了L-精氨酸向氨和L-硫氨酸的水解转化，这是能量产生的L-精氨酸降解途径的一部分。催化催化的化学机制涉及初始形成和随后的cys-烷基硫硫龙离子中间体的水解。铜绿假单胞菌的结构（L-精氨酸）复合物指导了可能与ADI反应的精氨酸类似物的设计，以在催化周转期间形成无活性的加合物。这样的候选者是L-甲烷，其中L-精氨酸的N-甲基烯被N-O取代。该物质被证明是慢慢产生的O-ureido-l-hoserine。 L-氯烷抑制铜绿假单胞菌ADI抑制的深度动力学和质谱分析表明，遵循了两种竞争途径，该途径是在Cys-碱基离子中间的分支。一条途径导致通过反应性硫龙中间体直接形成O- ureido-l-hoserine。其他途径导致酶的一种不活跃形式，该酶通过化学模型和质谱研究表明是Cys-Alkylisothiourea加合物。这种加合物经历了缓慢的水解，形成O- ureido-l-hoserine并再生酶。相比之下，动力学和质谱研究表明，在L-甲烷与熟食芽孢杆菌的反应中形成的Cys-烷基硫硫龙离子中间体在产物形成途径（O- ureido-l-l-ho-ho-homoserine and Free酶和灭活途径）之间的反应中形成了cys-烷基硫代基因研究。这会导致稳定的Cys-烷基硫代加合物加合物。检查了来自Escherichia大肠杆菌，Burkholderia Mallei和Giardia Intestinalis的ADI，以证明L-甲烷慢慢底物抑制的通用性用铜绿假单胞菌，E大肠杆菌，B。Mallei和G.观察到的抑制作用。 Intestinalis adis。
Arginine deiminase (ADI) catalyzes the hydrolytic conversion Of L-arginine to ammonia and L-Citrulline as part of the energy-producing L-arginine degradation pathway. The chemical mechanism for ADI catalysis involves initial formation and subsequent hydrolysis of a Cys-alkylthiouronium ion intermediate. The structure of the Pseudomonas aeruginosa ADI-(L-arginine) complex guided the design of arginine analogs that might react with the ADIS to form inactive covalent adducts during catalytic turnover. One such candidate is L-canavanine, in which an N-methylene of L-arginine is replaced by an N-O. This substance was shown to be a slow substrate-producing O-ureido-L-homoserine. An in depth kinetic and mass spectrometric analysis of P. aeruginosa ADI inhibition by L-canavanine showed that two competing pathways are followed that branch at the Cys-alkylthiouronium ion intermediate. One pathway leads to direct formation of O-ureido-L-homoserine via a reactive thiouronium intermediate. The other pathway leads to an inactive form of the enzyme, which was shown by chemical model and mass spectrometric studies to be a Cys-alkylisothiourea adduct. This adduct undergoes slow hydrolysis to form O-ureidO-L-homoserine and regenerated enzyme. In contrast, kinetic and mass spectrometric investigations demonstrate that the Cys-alkylthiouronium ion intermediate formed in the reaction Of L-canavanine with Bacillus cereus ADI partitions between the product forming pathway (O-ureido-L-homoserine and free enzyme) and an inactivation pathway that leads to a stable Cys-alkylthiocarbamate adduct. The ADIS from Escherichia coli, Burkholderia mallei, and Giardia intestinalis were examined in order to demonstrate the generality of the L-canavanine slow substrate inhibition and to distinguish the kinetic behavior that defines the irreversible inhibition observed with the B. cereus ADI from the time controlled inhibition observed with the P. aeruginosa, E coli, B. mallei, and G. intestinalis ADIS.