Structural Investigation of Enzymes that Utilize Cobalamin and AdoMet Cofactors

描述（由申请人提供）：在过去的几十年里，来自植物、动物或微生物的天然产物或化合物极大地激发了药物的发现和开发。由于自然界的化合物比现有的人造药物更复杂和结构独特，因此理解用于工程化这些分子的自然化学是基于天然产物的药物发现的重要里程碑。许多医学上重要的天然产物生物合成途径利用酶的化学性质，这些酶曾被认为是S-腺苷甲硫氨酸（S-adenosylmethionine，SNMet）自由基超家族的一个微小亚群的一部分。今天，这个亚组，钴胺素依赖性类的C3 Met自由基酶包含2，685个独特的成员，并且可以在重要化合物的生物合成途径中发现，包括磷霉素，福替米星A和硫链丝菌素A抗生素，以及抗病毒剂氧杂环辛A。尽管有大量的酶被确定属于这个亚组，而且它们催化的反应很重要，但关于它们如何发挥作用的机制细节却知之甚少。机制研究目前阻碍了这类酶的结构信息的缺乏和缺乏以前的特点，共享一个类似的序列，并使用相同的辅因子组合的酶。因此，本提案的目的是确定X-Met自由基超家族钴胺素依赖亚组成员的X射线晶体结构。这些结果将有助于设计未来的机制研究，并提供一个框架，了解协调和相互作用的维生素B1 Met和钴胺素辅因子在执行自然的化学。

DESCRIPTION (provided by applicant): Over the past few decades, natural products, or chemical compounds derived from plants, animals, or microbes have greatly inspired drug discovery and development. As nature's compounds are more complex and architecturally unique than available man-made drugs, understanding nature's chemistry for engineering these molecules represents a significant milestone in natural product based drug discovery. Many medically important natural product biosynthetic pathways utilize the chemistry of enzymes that were once thought to be part of a tiny subgroup of the S-adenosylmethionine (AdoMet) radical superfamily. Today, this subgroup, the cobalamin-dependent class of AdoMet radical enzymes contains 2,685 unique members and can be found in the biosynthetic pathways of important compounds including fosfomycin, fortimicin A, and thiostrepton A antibiotics, and the anti-viral agent oxetanocin A. Despite the wealth of enzymes identified as belonging to this subgroup and the importance of the reactions they catalyze, there are few mechanistic details known about how they function. Mechanistic studies are currently impeded by a lack of structural information for this class of enzymes and lack of previously characterized enzymes that share a similar sequence and use the same combination of cofactors. Therefore, the objective of this proposal is to determine X-ray crystal structures for members of the cobalamin-dependent subgroup of the AdoMet radical superfamily. These results will aid in the design of future mechanistic studies and provide a framework for understanding the coordination and interplay of AdoMet and cobalamin cofactors in performing nature's chemistry.