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PMID: 17932071 Published · ppublish English Journal Article Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't

Conserved amino acids in each subunit of the heteroligomeric tRNA m1A58 Mtase from Saccharomyces cerevisiae contribute to tRNA binding.

Nucleic acids research ·Vol. 35 ·No. 20 ·2007-00-00 ·Pages 6808-19

Ozanick SG, Bujnicki JM, Sem DS, Anderson JT

Abstract

In Saccharomyces cerevisiae, a two-subunit methyltransferase (Mtase) encoded by the essential genes TRM6 and TRM61 is responsible for the formation of 1-methyladenosine, a modified nucleoside found at position 58 in tRNA that is critical for the stability of tRNA(Met)i The crystal structure of the homotetrameric m1A58 tRNA Mtase from Mycobacterium tuberculosis, TrmI, has been solved and was used as a template to build a model of the yeast m1A58 tRNA Mtase heterotetramer. We altered amino acids in TRM6 and TRM61 that were predicted to be important for the stability of the heteroligomer based on this model. Yeast strains expressing trm6 and trm61 mutants exhibited growth phenotypes indicative of reduced m1A formation. In addition, recombinant mutant enzymes had reduced in vitro Mtase activity. We demonstrate that the mutations introduced do not prevent heteroligomer formation and do not disrupt binding of the cofactor S-adenosyl-L-methionine. Instead, amino acid substitutions in either Trm6p or Trm61p destroy the ability of the yeast m1A58 tRNA Mtase to bind tRNA(Met)i, indicating that each subunit contributes to tRNA binding and suggesting a structural alteration of the substrate-binding pocket occurs when these mutations are present.

MeSH Terms
Amino Acid Sequence Amino Acid Substitution Eukaryotic Initiation Factor-3/chemistry,metabolism Molecular Sequence Data Mutation RNA, Fungal/metabolism RNA, Transfer, Met/metabolism Saccharomyces cerevisiae/enzymology,genetics Saccharomyces cerevisiae Proteins/chemistry,metabolism Structure-Activity Relationship tRNA Methyltransferases/chemistry,genetics,metabolism
Chemicals
Eukaryotic Initiation Factor-3 RNA, Fungal RNA, Transfer, Met Saccharomyces cerevisiae Proteins Gcd10 protein, S cerevisiae tRNA Methyltransferases GCD14 protein, S cerevisiae
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Ozanick Sarah G
Department of Biological Sciences, Marquette University, P.O. Box 1881, Milwaukee, WI 53201, USA.
Bujnicki Janusz M
Sem Daniel S
Anderson James T
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Article Info
Journal
Nucleic acids research
Abbr.
Nucleic Acids Res
ISSN
1362-4962
DOI
10.1093/nar/gkm574
Published
2007-00-00
Epub
2007-00-10
Pages
6808-19
Language
English
Region
England
NLM ID
0411011
PMCID
PMC2175304
Subset
IM
Grants
NIGMS NIH HHS · R01GM069949 · United States
NCRR NIH HHS · S10 RR019012 · United States
NIGMS NIH HHS · R01 GM069949 · United States
NIGMS NIH HHS · R15 GM066791 · United States
NCRR NIH HHS · S10RR019012 · United States
NIGMS NIH HHS · R15GM066791-01 · United States
PHS HHS · P200A030199-05 · United States
External Links
PubMed source DOI: 10.1093/nar/gkm574 PMC Full Text View journal details
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