title: Conservation of structure and mechanism by Trm5 enzymes.
abstract: Enzymes of the Trm5 family catalyze methyl transfer from S-adenosyl methionine (AdoMet) to the N¹ of G37 to synthesize m¹ G37-tRNA as a critical determinant to prevent ribosome frameshift errors. Trm5 is specific to eukaryotes and archaea, and it is unrelated in evolution from the bacterial counterpart TrmD, which is a leading anti-bacterial target. The successful targeting of TrmD requires detailed information on Trm5 to avoid cross-species inhibition. However, most information on Trm5 is derived from studies of the archaeal enzyme Methanococcus jannaschii (MjTrm5), whereas little information is available for eukaryotic enzymes. Here we use human Trm5 (Homo sapiens; HsTrm5) as an example of eukaryotic enzymes and demonstrate that it has retained key features of catalytic properties of the archaeal MjTrm5, including the involvement of a general base to mediate one proton transfer. We also address the protease sensitivity of the human enzyme upon expression in bacteria. Using the tRNA-bound crystal structure of the archaeal enzyme as a model, we have identified a single substitution in the human enzyme that improves resistance to proteolysis. These results establish conservation in both the catalytic mechanism and overall structure of Trm5 between evolutionarily distant eukaryotic and archaeal species and validate the crystal structure of the archaeal enzyme as a useful model for studies of the human enzyme.
authors.affiliation: Department of Biochemistry and Molecular Biology, Thomas Jefferson University, Philadelphia, Pennsylvania 19107, USA.
authors.forename: Thomas
authors.initials: T
authors.lastname: Christian
authors.forename: Howard
authors.initials: H
authors.lastname: Gamper
authors.forename: Ya-Ming
authors.initials: YM
authors.lastname: Hou
mesh_terms: D019843:Archaeal Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D002384:Catalysis; D020134:Catalytic Domain; D006801:Humans; D007700:Kinetics; D017017:Methanococcus / Q000201:enzymology; D008745:Methylation; D008958:Models, Molecular; D011487:Protein Conformation; D012343:RNA, Transfer / Q000737:chemistry; D012436:S-Adenosylmethionine / Q000737:chemistry / Q000378:metabolism; D013379:Substrate Specificity; D012359:tRNA Methyltransferases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
chemical_list: D019843:Archaeal Proteins; D012436:S-Adenosylmethionine; D012343:RNA, Transfer; D012359:tRNA Methyltransferases; C489294:TRMT5 protein, human
keywords: S-adenosyl methionine; burst kinetics; m1G37-tRNA; pH-activity profile; structure-guided mutagenesis