Allen Kylie D, White Robert H
Department of Chemistry and Biochemistry, Gonzaga University, Spokane, WA, United States.
Virginia Polytechnic Institute and State University, Blacksburg, VA, United States.
Methods Enzymol. 2018;606:461-483. doi: 10.1016/bs.mie.2018.04.015. Epub 2018 May 10.
Methanogenic archaea represent a source of unique and fascinating anaerobic biochemistry that includes the involvement of many radical S-adenosyl-l-methionine (SAM) enzymes, some of which have well-established functions, while the majority have currently unknown or only partially understood functions. Here, we describe our strategy for the identification of the radical SAM enzyme that catalyzes the two methylation reactions in methanopterin biosynthesis in Methanocaldococcus jannaschii. Additionally, we describe the similar strategy carried out for the identification of the two radical SAM enzymes required for the biosynthesis of the 7,8-didemethyl-8-hydroxy-5-deazariboflavin (F) moiety of coenzyme F in M. jannaschii. This approach can be employed for future functional identification of radical SAM enzymes with currently unknown functions.
产甲烷古菌代表了独特且引人入胜的厌氧生物化学来源，其中包括许多自由基S-腺苷-L-甲硫氨酸（SAM）酶的参与，其中一些具有已明确的功能，而大多数目前功能未知或仅部分被理解。在此，我们描述了鉴定催化詹氏产甲烷球菌中喋呤生物合成中两个甲基化反应的自由基SAM酶的策略。此外，我们描述了为鉴定詹氏产甲烷球菌中辅酶F的7,8-二去甲基-8-羟基-5-脱氮核黄素（F）部分生物合成所需的两种自由基SAM酶而实施的类似策略。这种方法可用于未来对目前功能未知的自由基SAM酶进行功能鉴定。
