Evidence for nucleophilic catalysis in the aromatic substitution reaction catalyzed by (4-chlorobenzoyl)coenzyme A dehalogenase.
Evidence for nucleophilic catalysis in the aromatic substitution reaction catalyzed by (4-chlorobenzoyl)coenzyme A dehalogenase.
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(4-氯苯甲酰基)辅酶 A 脱卤酶催化的芳香族取代反应中亲核催化的证据。
DOI：
10.1021/bi00194a018
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发表时间：
1994
期刊：
影响因子：
2.9
通讯作者：
Dunaway-Mariano,D
中科院分区：
文献类型：
--
作者：
Yang,G;Liang,PH;Dunaway-Mariano,D
摘要：（4-氯苯甲酰）辅酶A脱卤酶催化（4-氯苯甲酰）辅酶A （4-CBA-CoA）水解脱卤为（4-羟基苯甲酰）辅酶A （4-HBA-CoA）。快速淬灭技术与[14C]-4-CBA-CoA一起用于测试催化过程中共价酶中间体的形成。在脱卤酶（75µ）过量2倍的情况下，测定[14C]-4-CBA-CoA（37µ）的消耗速率在k= 6.5 s™* 1下进行，与含有共价结合放射性标记物的酶中间体的形成一致。放射性标记酶在100 ms时达到最大值，相当于起始[14C]-4-CBA-CoA的27%，然后下降。在转化过程中观察到的放射性标记酶的形成和消耗动力学与它在整个反应中的中间作用是一致的。在98% 18g富集的水中进行单次翻转反应，在苯甲酰环C (4)-OH上产生73-75% 160和27-25% 180的4- hba - coa。相比之下，在93%的H2180中进行多次翻转反应，产生的4- hba - coa在C (4)-OH上标记为89% 180和11% 160。这些结果被解释为在4-CBA-CoA水解脱氯过程中在脱卤酶活性位点形成芳基酶的证据。（4-氯苯甲酰）辅酶A脱卤酶催化（4-氯苯甲酰）辅酶A (4-CBA-CoA) 1水解脱卤为（4-羟基苯甲酰）辅酶A （4-HBA-CoA）。该酶已在许多土壤细菌菌株中被发现，它与4-CBA: CoA连接酶和4-HBA-CoA硫酯酶一起在4-CBA到4-HBA的转化途径中起作用（方案1）[最近的综述，见Dunaway-Mariano和Babbitt(1994)]。4-CBA途径连接邻位裂解途径（从而连接/3-酮自适应途径），从而形成了该芳香化合物完全矿化的途径。虽然4-CBA不是天然产物，但它是在situó中作为4-氯联苯氧化生物降解的副产物产生的，4-氯联苯是一种普遍存在的合成污染物（Higson, 1992; Abramowicz, 1990; Commandeur & Parsons, 1990）。因此，4-CBA- coa脱卤酶在进化尺度上可能是相对较新的，它是最近由4-CBA的存在引发的基因重组的产物。对环境诱导的基因重组机制的深入了解可以从重组基因的谱系中获得。为此，我们寻找了4-CBA-CoA脱卤酶的祖先。我们发现
Revised Manuscript Received May 11, 1994· abstract:(4-Chlorobenzoyl) coenzyme A dehalogenase catalyzes the hydrolytic dehalogenation of (4-chlorobenzoyl) coenzyme A (4-CBA-CoA) to (4-hydroxybenzoyl) coenzyme A (4-HBA-CoA). Rapid-quench techniques were used in conjunction with [14C]-4-CBA-CoA to test for the formation of a covalent enzyme intermediate during catalysis. The rate of [14C]-4-CBA-CoA (37 µ) consumption in the presence of a 2-fold excess of dehalogenase (75 µ) was determined to proceed at k= 6.5 s™* 1, coincident with the formation of an enzyme intermediate containing covalently bound radiolabel. The radiolabeled enzyme reached a maximum level at 100 ms, corresponding to 27% of the starting [14C]-4-CBA-CoA, before declining. The kinetics of formation and consumption of the radiolabeled enzyme observed during turnover are consistent with its intermediacy in the overall reaction. A single turnover reaction carried out in 98% 18G-enriched water produced 4-HBA-CoA with 73-75% 160 and 27-25% 180 at the benzoyl ring C (4)-OH. In contrast, a multiple turnover reaction carried out in 93% H2180 produced 4-HBA-CoA labeled at the C (4)-OH with 89% 180 and 11% 160. Theseresults were interpreted as evidence for formation of an aryl enzyme intermediateduring 4-CBA-CoA hydrolytic dechlorination in thedehalogenase active site.(4-Chlorobenzoyl) coenzyme A dehalogenase catalyzes the hydrolytic dehalogenation of (4-chlorobenzoyl) coenzyme A (4-CBA-CoA) 1 to (4-hydroxybenzoyl) coenzyme A (4-HBA-CoA). This enzyme has been discovered in a number of soil-dwelling bacterial strains where it functions in a 4-CBA to 4-HBA converting pathway (Scheme 1) along with 4-CBA: CoA ligase and 4-HBA-CoA thioesterase [for a recent review, see Dunaway-Mariano and Babbitt (1994)]. The 4-CBA pathway connects to the ortho cleavage pathway (and hence to the/3-keto adipatepathway), thereby forming a route for the complete mineralization of this aromatic compound. While 4-CBA is not known to be a natural product, it is generated in situó as a byproduct of the oxidative biodegradation of 4-chlorobiphenyl, a ubiquitous, synthetic pollutant (Higson, 1992; Abramowicz, 1990; Commandeur & Parsons, 1990). Thus, the 4-CBA-CoA dehalogenase may be relatively new on the evolution scale, the product of recent gene retooling triggered by thepresence of 4-CBA. Insight into the mechanism of environmentally induced gene retooling might be gained from the lineage of the restructured gene. For this purpose, we have searched for the ancestors of the 4-CBA-CoA dehalogenase. We discovered that this
影响因子：
2.9
作者：
BABBITT, PC;KENYON, GL;DUNAWAYMARIANO, D
通讯作者：
DUNAWAYMARIANO, D
微粒体和可溶性环氧化物水解酶是同一 C-X 键水解酶家族的成员。
影响因子：
4.1
作者：
Lacourciere,GM;Armstrong,RN
通讯作者：
Armstrong,RN