研究白喉酰胺生物合成中自由基S-腺苷甲硫氨酸酶的方法。
Methods for Studying the Radical SAM Enzymes in Diphthamide Biosynthesis.
作者信息
Dong Min, Zhang Yugang, Lin Hening
机构信息
Department of Chemistry and Chemical Biology, Cornell University, Ithaca, NY, United States.
Howard Hughes Medical Institute, Cornell University, Ithaca, NY, United States.
出版信息
Methods Enzymol. 2018;606:421-438. doi: 10.1016/bs.mie.2018.04.001.
Diphthamide is a unique posttranslational modification on translation elongation factor 2 (EF2) in archaea and eukaryotes. Biosynthesis of diphthamide was proposed to involve four steps. The first step is a CC bond forming reaction catalyzed by unique radical S-adenosylmethionine (SAM) enzymes. Classical radical SAM enzymes use SAM and [4Fe-4S] clusters to generate a 5'-deoxyadenynal radical and catalyze numerous reactions. Radical SAM enzymes in diphthamide biosynthesis cleave a different CS bond in SAM to generate a 3-amino-3-carboxypropyl radical and modify a histidine residue of substrate protein EF2. Here, we describe our investigations on these unique radical SAM enzymes, including the preparation, characterization, and activity assays we have developed.
双氢乳清酸酰胺是古细菌和真核生物中翻译延伸因子2（EF2）上一种独特的翻译后修饰。双氢乳清酸酰胺的生物合成被认为涉及四个步骤。第一步是由独特的自由基S-腺苷甲硫氨酸（SAM）酶催化的CC键形成反应。经典的自由基SAM酶利用SAM和[4Fe-4S]簇生成5'-脱氧腺苷自由基并催化众多反应。双氢乳清酸酰胺生物合成中的自由基SAM酶切割SAM中不同的CS键以生成3-氨基-3-羧丙基自由基，并修饰底物蛋白EF2的一个组氨酸残基。在此，我们描述了对这些独特的自由基SAM酶的研究，包括我们所开展的制备、表征及活性测定。
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