The Ambiguous Genetic Code of Methanogenic Archaea that Grow on Methylamines

Natural genetic code expansion is a phenomenon wherein an additional amino acid is encoded by a stop codon. These non-standard amino acids are beneficial as they facilitate novel biochemical reactions. However, code expansion leads to ambiguity at the recoded stop codon, which can either be read through or terminated. Pyrrolysine (Pyl) is encoded by the amber codon (TAG/UAG) and is widespread in archaea, where it is required for methylamine-mediated methanogenesis, an environmentally important metabolism. Mechanisms to conditionally suppress the amber stop codon for Pyl installation during protein synthesis have not been identified. Using the model methanogen,Methanosarcina acetivorans,we demonstrate that Pyl-encoding archaea maintain an ambiguous genetic code wherein UAG encodes dual meaning as stop and Pyl. Our data suggest that expression of Pyl biosynthesis and incorporation genes is tuned to the cellular demand for Pyl, which allows these archaea to navigate ambiguous stop decoding in response to environmental cues.

We thank all members of the Nayak lab for their valuable feedback and support. DDN acknowledges funding from the Searle Scholars Program sponsored by the Kinship Foundation, the Rose Hills Innovator Grant, the Beckman Young Investigator Award sponsored by the Arnold and Mabel Beckman Foundation, the Alfred P. Sloan Research Fellowship sponsored by the Sloan Foundation, the Simons Foundation Early Career Investigator in Marine Microbial Ecology and Evolution Award, and the Packard Fellowship in Science and Engineering sponsored by the David and Lucille Packard Foundation. DDN is a Chan-Zuckerberg Biohub – San Francisco Investigator. KES was supported in part by the NSF Graduate Research Fellowship Program (Fellow ID: 202299857). GLC was supported by the Miller Institute for Basic Research in Science, University of California Berkeley. PIP was supported by University of California Berkeley’s Molecular and Cell Biology (MCB) department and the National Institute of Health (NIH) Bridges to Baccalaureate Program. DDN, PHW, VJO acknowledge funding from the Department of Energy through project number S589706. The funders had no role in the conceptualization and writing of this manuscript or the decision to submit the work for publication.