Structural and functional characterization of Rv2966c protein reveals an RsmD-like methyltransferase from Mycobacterium tuberculosis and the role of its N-terminal domain in target recognition

J Biol Chem. 2011 Jun 3;286(22):19652-61. doi: 10.1074/jbc.M110.200428. Epub 2011 Apr 7.

Abstract

Nine of ten methylated nucleotides of Escherichia coli 16 S rRNA are conserved in Mycobacterium tuberculosis. All the 10 different methyltransferases are known in E. coli, whereas only TlyA and GidB have been identified in mycobacteria. Here we have identified Rv2966c of M. tuberculosis as an ortholog of RsmD protein of E. coli. We have shown that rv2966c can complement rsmD-deleted E. coli cells. Recombinant Rv2966c can use 30 S ribosomes purified from rsmD-deleted E. coli as substrate and methylate G966 of 16 S rRNA in vitro. Structure determination of the protein shows the protein to be a two-domain structure with a short hairpin domain at the N terminus and a C-terminal domain with the S-adenosylmethionine-MT-fold. We show that the N-terminal hairpin is a minimalist functional domain that helps Rv2966c in target recognition. Deletion of the N-terminal domain prevents binding to nucleic acid substrates, and the truncated protein fails to carry out the m(2)G966 methylation on 16 S rRNA. The N-terminal domain also binds DNA efficiently, a property that may be utilized under specific conditions of cellular growth.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Bacterial Proteins / chemistry*
  • Bacterial Proteins / genetics
  • Bacterial Proteins / metabolism
  • Histones / chemistry*
  • Histones / genetics
  • Histones / metabolism
  • Methylation
  • Mycobacterium tuberculosis / enzymology*
  • Mycobacterium tuberculosis / genetics
  • Protein Structure, Tertiary
  • RNA, Bacterial / chemistry
  • RNA, Bacterial / genetics
  • RNA, Bacterial / metabolism
  • RNA, Ribosomal, 16S / chemistry
  • RNA, Ribosomal, 16S / genetics
  • RNA, Ribosomal, 16S / metabolism
  • Structure-Activity Relationship
  • tRNA Methyltransferases / chemistry*
  • tRNA Methyltransferases / genetics
  • tRNA Methyltransferases / metabolism

Substances

  • Bacterial Proteins
  • Histones
  • HupB protein, Mycobacterium tuberculosis
  • RNA, Bacterial
  • RNA, Ribosomal, 16S
  • tRNA Methyltransferases

Associated data

  • PDB/3P9N