Purification and characterization of an RNA dodecamer sequence binding protein from mitochondria of Saccharomyces cerevisiae

Biochem Biophys Res Commun. 1999 Aug 11;261(3):740-5. doi: 10.1006/bbrc.1999.1085.

Abstract

Saccharomyces cerevisiae mitochondrial mRNAs terminate at their 3' ends with a conserved dodecamer sequence, 5'-AAUAA(U/C)AUUCUU-3'. We have identified a nuclear-encoded protein (DBP) which specifically binds to the dodecamer sequence and have purified it to apparent homogeneity by RNA affinity chromatography. DBP consists of a single polypeptide of 55 kDa and binds to its RNA substrate with a 1:1 stoichiometry. Scatchard analysis determines that K(d) is 0.93 nM for the canonical dodecamer sequence (5'-AAUAAUAUUCUU-3') and 0.46 nM for the only naturally occurring variant (5'-AAUAACAUUCUU-3') unique to oli1 gene. Based on the studies using mutant oligonucleotides, DBP appears to recognize primarily the nucleotide sequence of an RNA rather than its potential secondary structure.

MeSH terms

  • Base Sequence
  • Binding Sites
  • Mitochondria / chemistry*
  • Nucleic Acid Conformation
  • Oligonucleotides / chemistry
  • Oligonucleotides / metabolism
  • RNA / chemistry
  • RNA / metabolism
  • RNA-Binding Proteins / isolation & purification*
  • RNA-Binding Proteins / metabolism
  • Saccharomyces cerevisiae / ultrastructure*

Substances

  • Oligonucleotides
  • RNA-Binding Proteins
  • RNA