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首页> 外文期刊>Journal of bacteriology >Degradation of Aromatics and Chloroaromatics by Pseudomonas sp. Strain B13: Cloning, Characterization, and Analysis of Sequences Encoding 3-Oxoadipate:Succinyl-Coenzyme A (CoA) Transferase and 3-Oxoadipyl-CoA Thiolase
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Degradation of Aromatics and Chloroaromatics by Pseudomonas sp. Strain B13: Cloning, Characterization, and Analysis of Sequences Encoding 3-Oxoadipate:Succinyl-Coenzyme A (CoA) Transferase and 3-Oxoadipyl-CoA Thiolase

机译：假单胞菌（Pseudomonas sp。）降解芳香族和氯代芳香族。菌株B13：克隆，表征和编码3-氧代己二酸：琥珀酰辅酶A（CoA）转移酶和3-氧代己二酰-CoA硫醇酶的序列的分析

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摘要

3-Oxoadipate:succinyl-coenzyme A (CoA) transferase and 3-oxoadipyl-CoA thiolase carry out the ultimate steps in the conversion of benzoate and 3-chlorobenzoate to tricarboxylic acid cycle intermediates in bacteria utilizing the 3-oxoadipate pathway. This report describes the characterization of DNA fragments with the overall length of 5.9 kb from Pseudomonas sp. strain B13 that encode these enzymes. DNA sequence analysis revealed five open reading frames (ORFs) plus an incomplete one. ORF1, of unknown function, has a length of 414 bp. ORF2 (catI) encodes a polypeptide of 282 amino acids and starts at nucleotide 813. ORF3 (catJ) encodes a polypeptide of 260 amino acids and begins at nucleotide 1661. CatI and CatJ are the subunits of the 3-oxoadipate:succinyl-CoA transferase, whose activity was demonstrated when both genes were ligated into expression vector pET11a. ORF4, termed catF, codes for a protein of 401 amino acid residues with a predicted mass of 41,678 Da with 3-oxoadipyl-CoA thiolase activity. The last three ORFs seem to form an operon since they are oriented in the same direction and showed an overlapping of 1 bp between catI and catJ and of 4 bp between catJ and catF. Conserved functional groups important for the catalytic activity of CoA transferases and thiolases were identified in CatI, CatJ, and CatF. ORF5 (catD) encodes the 3-oxoadipate enol-lactone hydrolase. An incomplete ORF6 of 1,183 bp downstream of ORF5 and oriented in the opposite direction was found. The protein sequence deduced from ORF6 showed a putative AMP-binding domain signature.
机译：3-氧代己二酸：琥珀酰辅酶A（CoA）转移酶和3-氧代己二酰-CoA硫解酶在利用3-氧代己二酸途径将细菌中的苯甲酸酯和3-氯苯甲酸酯转化为三羧酸循环中间体的过程中执行了最终步骤。该报告描述了来自假单胞菌 sp。的全长5.9 kb的DNA片段的表征。编码这些酶的B13菌株。 DNA序列分析揭示了五个开放阅读框（ORF）和一个不完整的阅读框。功能未知的ORF1的长度为414 bp。 ORF2（ catI ）编码一个282个氨基酸的多肽，起始于核苷酸813。ORF3（ catJ ）编码一个260个氨基酸的多肽，起始于核苷酸1661。 CatJ是3-氧己二酸酯：琥珀酰-CoA转移酶的亚基，当两个基因都连接到表达载体pET11a中时，其活性得到了证实。称为 catF 的ORF4编码具有401个氨基酸残基的蛋白质，预计质量为41,678 Da，具有3-氧代己二酰辅酶A硫解酶的活性。后三个ORF似乎形成操纵子，因为它们的方向相同，并且在 catI 和 catJ 之间重叠1 bp，在之间重叠4 bp。 catJ 和 catF 。在CatI，CatJ和CatF中鉴定了对CoA转移酶和硫糖酶催化活性重要的保守官能团。 ORF5（ catD ）编码3-氧代己二酸烯醇内酯水解酶。发现一个不完整的ORF6，位于ORF5下游1,183 bp，方向相反。从ORF6推导的蛋白质序列显示出一个假定的AMP结合域签名。

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