Methanogenesis from tetramethylammonium and choline in Methanococcoides methylutens Q3c requires a nonpyrrolysine monomethylamine methyltransferase homolog

Jyoti Kashyap¹Tomislav Ticak²Adam J. Creighbaum²Khatri Rakhsha³Donald J Ferguson^2,4*

• ¹University of Idaho, Moscow, United States
• ²Miami University, Oxford, United States
• ³Wright State University Boonshoft School of Medicine, Dayton, United States
• ⁴Miami University - Hamilton Campus, Hamilton, United States

Methanogenesis is a key biological process contributing to both global climate change and renewable energy via methane. Methylotrophic methanogenesis is an important, but often underappreciated, component of this process. Recently, there has been an increasing interest in methylotrophic methanogenesis from quaternary amines such as glycine betaine, choline, and tetramethylammonium (QMA). QMA was the original quaternary amine identified as a direct substrate for methanogens and the components of its catabolic pathway were isolated from Methanococcoides methylutens NaT1. Loss of strain NaT1 and the absence of genomic sequencing leaves into question the identity of the genes responsible. We previously isolated a related strain of Methanococcoides methylutens called strain Q3c, capable of growth on the quaternary amines QMA and choline. We conducted proteomic analysis of strain Q3c in which we identified potential targets for QMA and choline metabolism, involving the methyltransferase, MtqB, and its cognate corrinoid protein, MtqC, because they were highly produced in QMA-grown and choline-grown cells differentially compared to trimethylamine grown cells and the 82.8% and 96.4%, identity of their N-termini, respectively, to MtqBC identified previously in NaT1. Surprisingly, MtqB was identified as a nonpyrrolysine monomethylamine methyltransferase (MtmB) homolog. This was validated via enzymatic assay and molecular co-docking of MtqB with QMA and choline along with its cognate partner MtqC, representing the first ever function for a non-Pyl MtmB demonstrated. The phylogeny of MtqB and MtqC showed unique clustering of these proteins compared to other homologs, suggesting these proteins may be important in our continued study of the evolution of Pyl-and non-Pyl methyltransferase systems.