Ferguson Donald J, Longstaff David G, Krzycki Joseph A
Department of Microbiology, University of Miami, Oxford, Ohio, USA.
Methods Enzymol. 2011;494:139-58. doi: 10.1016/B978-0-12-385112-3.00008-1.
The family Methanosarcinaceae has an expanded repertoire of growth substrates relative to most other methanogenic archaea. Various methylamines, methylated thiols, and methanol can serve as precursors to both methane and carbon dioxide. These compounds are mobilized into metabolism by methyltransferases that use the growth substrate to methylate a cognate corrinoid protein, which in turn is used as a substrate by a second methyltransferase to methylate Coenzyme M (CoM), forming methyl-SCoM, the precursor to both methane and carbon dioxide. Orthologs of the methyltransferases, as well as the small corrinoid proteins, are found in many archaeal and bacterial genomes. Some of these are homologs of the methylamine methyltransferases predicted to require pyrrolysine, an atypical genetically encoded amino acid, for synthesis. As a resource for the study of these sizable families of proteins, we describe here techniques our laboratories have used for the study of methanogen corrinoid-dependent methyltransferases, focusing especially on isolation and assay techniques useful for various activities of components of the methylamine- and methylthiol-dependent CoM methyltransferase systems.
相对于大多数其他产甲烷古菌，甲烷八叠球菌科具有更广泛的生长底物种类。各种甲胺、甲基化硫醇和甲醇都可以作为甲烷和二氧化碳的前体。这些化合物通过甲基转移酶进入代谢过程，甲基转移酶利用生长底物将同源类咕啉蛋白甲基化，而类咕啉蛋白又被第二种甲基转移酶用作底物来甲基化辅酶M（CoM），形成甲基-SCoM，它是甲烷和二氧化碳的前体。甲基转移酶以及小型类咕啉蛋白的直系同源物存在于许多古菌和细菌基因组中。其中一些是预计需要吡咯赖氨酸（一种非典型的遗传编码氨基酸）来合成的甲胺甲基转移酶的同源物。作为研究这些庞大蛋白质家族的资源，我们在此描述我们实验室用于研究产甲烷菌类咕啉依赖性甲基转移酶的技术，特别关注对甲胺和甲基硫醇依赖性CoM甲基转移酶系统各组分的各种活性有用的分离和检测技术。
