Lichstrahl Michael S, Townsend Craig A, Sinner Erica K
Department of Chemistry, The Johns Hopkins University 3400 N Charles St Baltimore Maryland USA
RSC Chem Biol. 2022 Jun 6;3(8):1028-1034. doi: 10.1039/d2cb00113f. eCollection 2022 Aug 3.
Complex carbapenems are important clinical antibiotics for difficult-to-treat infections. An essential step in the biosyntheses of these natural products is stereospecific methylation at C6 and subsequent alkylations by cobalamin-dependent radical SAM methylases such as TokK and ThnK. We have prepared isotopically labeled substrates in a stereospecific manner and found that both homologous enzymes selectively abstract the 6-pro- hydrogen, followed by methyl transfer to the opposite face to give the (6)-methyl carbapenam product proceeding, therefore, by inversion of absolute configuration at C6. These data clarify an unexpected ambiguity in the recently solved substrate-bound crystal structure of TokK and have led to a stereochemically complete mechanistic proposal for both TokK and ThnK.
复杂碳青霉烯类是用于治疗难治性感染的重要临床抗生素。这些天然产物生物合成中的一个关键步骤是在C6位进行立体特异性甲基化，随后由钴胺素依赖性自由基SAM甲基转移酶（如TokK和ThnK）进行烷基化。我们以立体特异性方式制备了同位素标记的底物，发现这两种同源酶都选择性地提取6-前氢，然后甲基转移到相反的面，从而生成(6)-甲基碳青霉烯产物，因此，C6位的绝对构型发生了翻转。这些数据澄清了TokK最近解析的底物结合晶体结构中一个意想不到的模糊之处，并为TokK和ThnK提出了一个立体化学完整的机理方案。
