Department of Chemistry, Stanford University, Stanford, California 94305, USA.
Biochemistry. 2010 Feb 16;49(6):1176-82. doi: 10.1021/bi901772w.
Isopenicillin N synthase (IPNS) can have both oxidase and oxygenase activity depending on the substrate. For the native substrate, ACV, oxidase activity exists; however, for the substrate analogue ACOV, which lacks an amide nitrogen, IPNS exhibits oxygenase activity. The potential energy surfaces for the O-O bond elongation and cleavage were calculated for three different reactions: homolytic cleavage via traditional Fenton chemistry, heterolytic cleavage, and nucleophilic attack. These surfaces show that the hydroperoxide-ferrous intermediate, formed by O(2)-activated H atom abstraction from the substrate, can exploit different reaction pathways and that interactions with the substrate govern the pathway. The hydrogen bonds from hydroperoxide to the amide nitrogen of ACV polarize the sigma* orbital of the peroxide toward the proximal oxygen, facilitating heterolytic cleavage. For the substrate analogue ACOV, this hydrogen bond is no longer present, leading to nucleophilic attack on the substrate intermediate C-S bond. After cleavage of the hydroperoxide, the two reaction pathways proceed with minimal barriers, resulting in the closure of the beta-lactam ring for the oxidase activity (ACV) or formation of the thiocarboxylate for oxygenase activity (ACOV).
异青霉素 N 合酶 (IPNS) 可以根据底物表现出氧化酶和加氧酶活性。对于天然底物 ACV，存在氧化酶活性；然而，对于缺乏酰胺氮的底物类似物 ACOV，IPNS 表现出加氧酶活性。计算了三种不同反应的 O-O 键延长和断裂的势能面：通过传统的芬顿化学进行均裂断裂、异裂断裂和亲核攻击。这些表面表明，由 O(2)-从底物中激活的 H 原子夺取形成的过氧化物-亚铁中间物可以利用不同的反应途径，并且与底物的相互作用控制途径。过氧化物中的氢键将酰胺氮的 sigma*轨道极化到过氧化物的近端氧，促进异裂断裂。对于底物类似物 ACOV，这种氢键不再存在，导致亲核攻击底物中间的 C-S 键。过氧化物断裂后，两条反应途径以最小的能垒进行，导致氧化酶活性（ACV）中β-内酰胺环闭合或加氧酶活性（ACOV）中硫代羧酸酯形成。
