The highly conserved KEOPS/EKC complex is essential for a universal tRNA modification, t6A

EMBO J. 2011 Mar 2;30(5):873-81. doi: 10.1038/emboj.2010.343. Epub 2010 Dec 24.

Abstract

The highly conserved Kinase, Endopeptidase and Other Proteins of small Size (KEOPS)/Endopeptidase-like and Kinase associated to transcribed Chromatin (EKC) protein complex has been implicated in transcription, telomere maintenance and chromosome segregation, but its exact function remains unknown. The complex consists of five proteins, Kinase-Associated Endopeptidase (Kae1), a highly conserved protein present in bacteria, archaea and eukaryotes, a kinase (Bud32) and three additional small polypeptides. We showed that the complex is required for a universal tRNA modification, threonyl carbamoyl adenosine (t6A), found in all tRNAs that pair with ANN codons in mRNA. We also showed that the bacterial ortholog of Kae1, YgjD, is required for t6A modification of Escherichia coli tRNAs. The ATPase activity of Kae1 and the kinase activity of Bud32 are required for the modification. The yeast protein Sua5 has been reported previously to be required for t6A synthesis. Using yeast extracts, we established an in vitro system for the synthesis of t6A that requires Sua5, Kae1, threonine, bicarbonate and ATP. It remains to be determined whether all reported defects of KEOPS/EKC mutants can be attributed to the lack of t6A, or whether the complex has multiple functions.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Adenosine / analogs & derivatives*
  • Adenosine / metabolism
  • Amino Acid Sequence
  • Base Sequence
  • Metalloendopeptidases / genetics
  • Metalloendopeptidases / metabolism*
  • Molecular Sequence Data
  • Multiprotein Complexes
  • Nucleic Acid Conformation
  • Protein Biosynthesis
  • Protein-Serine-Threonine Kinases / genetics
  • Protein-Serine-Threonine Kinases / metabolism*
  • RNA, Transfer / chemistry*
  • RNA, Transfer / genetics
  • RNA, Transfer / metabolism
  • Saccharomyces cerevisiae / genetics*
  • Saccharomyces cerevisiae / growth & development
  • Saccharomyces cerevisiae / metabolism
  • Saccharomyces cerevisiae Proteins / genetics
  • Saccharomyces cerevisiae Proteins / metabolism*
  • Sequence Homology, Amino Acid
  • Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization

Substances

  • KEOPS complex, S cerevisiae
  • Multiprotein Complexes
  • Saccharomyces cerevisiae Proteins
  • N(6)-(N-threonylcarbonyl)adenosine
  • RNA, Transfer
  • BUD32 protein, S cerevisiae
  • Protein-Serine-Threonine Kinases
  • Kae1 protein, S cerevisiae
  • Metalloendopeptidases
  • Adenosine