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论文类型：期刊论文

发表时间：2018-12-27

发表刊物：JOURNAL OF PHYSICAL CHEMISTRY B

收录刊物：SCIE、Scopus

卷号：122

期号：51

页面范围：12241-12250

ISSN号：1520-6106

关键字：Carboxylation; Chemical activation; Enzymes; Fatty acids; Hydrides; Ligands; Stereochemistry; Stereoselectivity; Substrates, Activation barriers; Catalytic transformation; Experimental investigations; Fatty acid synthesis; Protein engineering; Stereoselective recognition; Structural knowledge; Transition state theories, Biochemical engineering

摘要：In protein engineering, investigations of catalytic cycle facilitate rational design of enzymes. In the present work, deeper analysis on the catalytic cycle of malate enzyme (EC 1.1.1.40), an enzyme involved in cancer metabolic and fatty acid synthesis, was performed. In substrate binding, stereoselective recognition of a substrate originates from distance and angle difference between two chiral substrates and Mn2+ as well as monodentate or coplanar ion reaction with Arg165. In catalytic transformation, the activation barrier for the hydride transfer of D-malate is 20.28 kcal/mol higher than that for L-malate. The activation barrier for beta-decarboxylation of oxaloacetate is about 4.59 kcal/mol higher than the activation barrier for the hydride transfer of L-malate. The effective activation barrier is 16.44 kcal/mol, which is in close agreement with the value derived from the application of transition-state theory and the Eyring equation to k(cat). In ligand release, L/D-malate needs to overcome a higher barrier than pyruvate to break all bonds in parallel and then to escape from the binding pocket. Leu167 and Asn421 comprise a swinging gate to control the product release. The more open gate is possibly required in the direction of pyruvate to L-malate. Our studies are focused on extending structural knowledge regarding the malate enzyme and provided a powerful strategy for future experimental investigations.