Institute of Biological Sciences Faculty of Science University of Malaya Kuala Lumpur Malaysia.
Department of Biochemistry College of Health Sciences Bayero University Kano Nigeria.
FEBS Open Bio. 2019 Jan 30;9(3):408-419. doi: 10.1002/2211-5463.12405. eCollection 2019 Mar.
A glutathione -transferase (GST) with a potential dehalogenation function against various organochlorine substrates was identified from a polychlorobiphenyl (PCB)-degrading organism, sp. KKS102. A homolog of the gene (biphenyl upper pathway K), named , was cloned, purified and biochemically characterized. Bioinformatic analysis indicated several conserved amino acids that participated in the catalytic activity of the enzyme, and site-directed mutagenesis of these conserved amino acids revealed their importance in the enzyme's catalytic activity. The wild-type and mutant (C10F, K107T and A180P) recombinant proteins displayed wider substrate specificity. The wild-type recombinant GST reacted towards 1-chloro-2,4-dinitrobenzene (CDNB), ethacrynic acid, hydrogen peroxide and cumene hydroperoxide. The mutated recombinant proteins, however, showed significant variation in specific activities towards the substrates. A combination of a molecular docking study and a chloride ion detection assay showed potential interaction with and a dechlorination function against 2-, 3- and 4-chlorobenzoates (metabolites generated during PCB biodegradation) in addition to some organochlorine pesticides (dichlorodiphenyltrichloroethane, endosulfan and permethrin). It was demonstrated that the behavior of the dechlorinating activities varied among the wild-type and mutant recombinant proteins. Kinetic studies (using CDNB and glutathione) showed that the kinetic parameters , , and / were all affected by the mutations. While C10F and A180P mutants displayed an increase in GST activity and the dechlorination function of the enzyme, the K107T mutant displayed variable results, suggesting a functional role of Lys107 in determining substrate specificity of the enzyme. These results demonstrated that the enzyme should be valuable in the bioremediation of metabolites generated during PCB biodegradation.
从多氯联苯（PCB）降解菌 sp. KKS102 中鉴定出一种具有潜在脱卤功能的谷胱甘肽转移酶（GST），可作用于各种有机氯底物。该基因的同源物（联苯上途径 K）被命名为 ，被克隆、纯化并进行了生化特性分析。生物信息学分析表明，该酶的催化活性涉及几个保守氨基酸，对这些保守氨基酸进行定点突变揭示了它们在酶的催化活性中的重要性。野生型和突变型（C10F、K107T 和 A180P）重组蛋白显示出更广泛的底物特异性。野生型重组 GST 可与 1-氯-2,4-二硝基苯（CDNB）、乙叉基脲、过氧化氢和cumene 氢过氧化物反应。然而，突变型重组蛋白对这些底物的比活性表现出显著差异。分子对接研究和氯离子检测实验表明，该酶除了与一些有机氯农药（滴滴涕、硫丹和氯菊酯）外，还可能与 2-、3-和 4-氯苯甲酸（PCB 生物降解过程中的代谢物）发生相互作用并具有脱卤功能。结果表明，野生型和突变型重组蛋白的脱卤活性行为存在差异。动力学研究（使用 CDNB 和谷胱甘肽）表明，突变影响了动力学参数 、 、 和 /。虽然 C10F 和 A180P 突变体显示 GST 活性和酶的脱卤功能增加，但 K107T 突变体的结果则不同，表明 Lys107 在确定酶的底物特异性方面具有功能作用。这些结果表明，该酶在 PCB 生物降解过程中产生的代谢物的生物修复中具有潜在的应用价值。
