Structural and functional insights into the molecular mechanism of rRNA m6A methyltransferase RlmJ

Avinash S. Punekar, Josefine Liljeruhm, Tyson R. Shepherd, Anthony C. Forster, Maria Selmer (Lead / Corresponding author)

Research output: Contribution to journalArticle

12 Citations (Scopus)

Abstract

RlmJ catalyzes the m6A2030 methylation of 23S rRNA during ribosome biogenesis in Escherichia coli. Here, we present crystal structures of RlmJ in apo form, in complex with the cofactor S-adenosyl-methionine and in complex with S-adenosyl-homocysteine plus the substrate analogue adenosine monophosphate (AMP). RlmJ displays a variant of the Rossmann-like methyltransferase (MTase) fold with an inserted helical subdomain. Binding of cofactor and substrate induces a large shift of the N-terminal motif X tail to make it cover the cofactor binding site and trigger active-site changes in motifs IV and VIII. Adenosine monophosphate binds in a partly accommodated state with the target N6 atom 7 Å away from the sulphur of AdoHcy. The active site of RlmJ with motif IV sequence 164DPPY167 is more similar to DNA m6A MTases than to RNA m62A MTases, and structural comparison suggests that RlmJ binds its substrate base similarly to DNA MTases T4Dam and M.TaqI. RlmJ methylates in vitro transcribed 23S rRNA, as well as a minimal substrate corresponding to helix 72, demonstrating independence of previous modifications and tertiary interactions in the RNA substrate. RlmJ displays specificity for adenosine, and mutagenesis experiments demonstrate the critical roles of residues Y4, H6, K18 and D164 in methyl transfer.

Original languageEnglish
Pages (from-to)9537-9548
Number of pages12
JournalNucleic Acids Research
Volume41
Issue number20
DOIs
Publication statusPublished - Nov 2013

Keywords

  • Adenine/analogs & derivatives
  • Adenosine Monophosphate/chemistry
  • Amino Acid Sequence
  • Binding Sites
  • Catalytic Domain
  • Coenzymes/chemistry
  • Escherichia coli/enzymology
  • Escherichia coli Proteins/chemistry
  • Methyltransferases/chemistry
  • Models, Molecular
  • Molecular Sequence Data
  • RNA, Ribosomal, 23S/chemistry
  • S-Adenosylhomocysteine/chemistry
  • S-Adenosylmethionine/chemistry
  • Sequence Alignment

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