Hindawi Publishing Corporation Archaea Volume 2013, Article ID 185250, 7 pages http://dx.doi.org/10.1155/2013/185250 Research Article Genetic Confirmation of the Role of Sulfopyruvate Decarboxylase in Coenzyme M Biosynthesis in Methanococcus maripaludis Felipe Sarmiento, Courtney K. Ellison, and William B. Whitman Department of Microbiology, University of Georgia, 541 Biological Science Building, Athens, GA 30602-2605, USA Correspondence should be addressed to William B. Whitman; whitman@uga.edu Received 22 May 2013; Accepted 8 August 2013 Academic Editor: Stefan Spring Copyright © 2013 Felipe Sarmiento et al. Tis is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Coenzyme M is an essential coenzyme for methanogenesis. Te proposed biosynthetic pathway consists of fve steps, of which the fourth step is catalyzed by sulfopyruvate decarboxylase (ComDE). Disruption of the gene comE by transposon mutagenesis resulted in a partial coenzyme M auxotroph, which grew poorly in the absence of coenzyme M and retained less than 3% of the wild type level of coenzyme M biosynthesis. Upon coenzyme M addition, normal growth of the mutant was restored. Moreover, complementation of the mutation with the wild type comE gene in trans restored full growth in the absence of coenzyme M. Tese results confrm that ComE plays an important role in coenzyme M biosynthesis. Te inability to yield a complete CoM auxotroph suggests that either the transposon insertion failed to completely inactivate the gene or M. maripaludis possesses a promiscuous activity that partially complemented the mutation. 1. Introduction Hydrogenotrophic methanogens, such as Methanococcus maripaludis, possess a specialized metabolism. In a process known as methanogenesis, they reduce CO 2 to CH 4 using H 2 or formate as the electron donor [1]. While other methanoar- chaea can use acetate, methylamine, and other methyl-group- containing compounds [1], coenzyme M (CoM), the smallest known organic cofactor, plays a key role as the last methyl carrier in all methanogens [2]. Tus, methane is formed upon the reduction of methyl-CoM with coenzyme B (CoB) as an electron donor by the methyl-CoM reductase. Te oxidation of CoB yields a heterodisulfde with CoM (CoM-S-S-CoB), which is reduced to regenerate the thiols by heterodisulfde reductase (Hdr) [3]. Without coenzyme M being present to complete the biosynthesis of methane, the organism is unable to produce the necessary energy for growth. Te biosynthetic pathway of coenzyme M (CoM) in Methanocaldococcus jannaschii, an organism closely related to M. maripaludis, is proposed to proceed in fve steps. Four enzymes involved in the biosynthesis of CoM have been biochemically characterized [7–10]. In addition, the genes encoding these enzymes have been identifed in diverse methanogens, including M. maripaludis. Te proposed path- way for the biosynthesis of CoM starts with the sulfonation of PEP by a phosphoenolpyruvate sulfotransferase (ComA). Ten, a phosphosulfolactate phosphatase (ComB) hydrolyses phosphosulfolactate, and a dehydrogenase (ComC) oxidizes the (R)-sulfolactate intermediate to form sulfopyruvate. In the fourth step, a sulfopyruvate decarboxylase (ComDE) catalyzes the decarboxylation of sulfopyruvate to form sul- foacetaldehyde [10]. For the fnal postulated step of CoM biosynthesis, sulfoacetaldehyde is reductively thiolated to form coenzyme M. However, this enzyme has not yet been identifed. In addition, the genomes of Methanosarcina species lack the genes for the frst three steps of the pathway, suggesting that an alternative pathway for CoM biosynthesis exists [2]. Lastly, none of the steps of this pathway have been confrmed in vivo by mutagenesis of the proposed genes. M. maripaludis may have the ability to take up coenzyme M from the medium. Previous studies have identifed an energy-dependent transport system for coenzyme M within