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首页> 外文期刊>Applied Microbiology >2-Haloacrylate Hydratase, a New Class of Flavoenzyme That Catalyzes the Addition of Water to the Substrate for Dehalogenation
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2-Haloacrylate Hydratase, a New Class of Flavoenzyme That Catalyzes the Addition of Water to the Substrate for Dehalogenation

机译：2-卤代丙烯酸水合酶，一类新型的黄素酶，催化水加到基质上进行脱卤

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Enzymes catalyzing the conversion of organohalogen compounds are useful in the chemical industry and environmental technology. Here we report the occurrence of a new reduced flavin adenine dinucleotide (FAD) (FADH_(2))-dependent enzyme that catalyzes the removal of a halogen atom from an unsaturated aliphatic organohalogen compound by the addition of a water molecule to the substrate. A soil bacterium, Pseudomonas sp. strain YL, inducibly produced a protein named Caa67_(YL) when the cells were grown on 2-chloroacrylate (2-CAA). The caa67 _(YL) gene encoded a protein of 547 amino acid residues ( M _(r) of 59,301), which shared weak but significant sequence similarity with various flavoenzymes and contained a nucleotide-binding motif. We found that 2-CAA is converted into pyruvate when the reaction was carried out with purified Caa67_(YL) in the presence of FAD and a reducing agent [NAD(P)H or sodium dithionite] under anaerobic conditions. The reducing agent was not stoichiometrically consumed during this reaction, suggesting that FADH_(2) is conserved by regeneration in the catalytic cycle. When the reaction was carried out in the presence of H_(2)~(18)O, [~(18)O]pyruvate was produced. These results indicate that Caa67_(YL) catalyzes the hydration of 2-CAA to form 2-chloro-2-hydroxypropionate, which is chemically unstable and probably spontaneously dechlorinated to form pyruvate. 2-Bromoacrylate, but not other 2-CAA analogs such as acrylate and methacrylate, served as the substrate of Caa67_(YL). Thus, we named this new enzyme 2-haloacrylate hydratase. The enzyme is very unusual in that it requires the reduced form of FAD for hydration, which involves no net change in the redox state of the coenzyme or substrate.
机译：催化有机卤素化合物转化的酶可用于化学工业和环境技术。在这里，我们报告新的还原黄素腺嘌呤二核苷酸（FAD）（FADH_（2））依赖酶的发生，该酶通过向底物中添加水分子来催化从不饱和脂肪族有机卤素化合物中去除卤素原子。土壤细菌，假单胞菌。当细胞在2-氯丙烯酸酯（2-CAA）上生长时，菌株YL可诱导产生名为Caa67_（YL）的蛋白质。 caa67_（YL）基因编码一个547个氨基酸残基的蛋白质（M_（r）为59,301），与各种黄素酶具有弱但显着的序列相似性，并包含一个核苷酸结合基序。我们发现，当在厌氧条件下，在FAD和还原剂[NAD（P）H或连二亚硫酸钠]存在下，用纯化的Caa67_（YL）进行反应时，2-CAA会转化为丙酮酸。在该反应过程中，还原剂没有以化学计量消耗，表明FADH_（2）通过催化循环中的再生而得以保存。当反应在H_（2）〜（18）O存在下进行时，生成[〜（18）O]丙酮酸盐。这些结果表明，Caa67_（YL）催化2-CAA的水合反应形成2-氯-2-羟基丙酸酯，该化合物化学性质不稳定，可能会自发脱氯形成丙酮酸。 2-溴丙烯酸酯，而不是其他2-CAA类似物（例如丙烯酸酯和甲基丙烯酸酯）用作Caa67_（YL）的底物。因此，我们将这种新酶命名为2-卤代丙烯酸酯水合酶。该酶是非常不寻常的，因为它需要还原形式的FAD来水合，这不涉及辅酶或底物的氧化还原状态的净变化。

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