A chimeric ubiquitin conjugating enzyme that combines the cell cycle properties of CDC34 (UBC3) and the DNA repair properties of RAD6 (UBC2): implications for the structure, function and evolution of the E2s

EMBO J. 1992 Aug;11(8):3091-8.

Abstract

The CDC34 (UBC3) protein from Saccharomyces cerevisiae has a 125 residue tail that contains a polyacidic region flanked on either side by sequences of mixed composition. We show that although a catalytic domain is essential for CDC34 activity, a major cell cycle determinant of this enzyme is found within a 74 residue segment of the tail that does not include the polyacidic stretch or downstream sequences. Transposition of the CDC34 tail onto the catalytic domain of a functionally unrelated E2 such as RAD6 (UBC2) results in a chimeric E2 that combines RAD6 and CDC34 activities within the same polypeptide. In addition to the tail, the cell cycle function exhibited by the chimera and CDC34 is probably dependent on a conserved region of the catalytic domain that is shared by both RAD6 and CDC34. Despite this similarity, the CDC34 catalytic domain cannot substitute for the DNA repair and growth functions of the RAD6 catalytic domain, indicating that although these domains are structurally related, sufficient differences exist to maintain their functional individuality. Expression of the CDC34 catalytic domain and tail as separate polypeptides are capable of only partial function; thus, while the tail displays autonomous structural characteristics, there is considerable advantage gained when both domains coexist within the same polypeptide. The ability of these and other derivatives to restore partial function to a cdc34 temperature-sensitive mutant but not to a disruption mutant suggests that interaction between two CDC34 polypeptides is a requirement of CDC34 activity. Based on this idea we propose a model that accounts for the initiating steps leading to multi-ubiquitin chain synthesis.(ABSTRACT TRUNCATED AT 250 WORDS)

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Anaphase-Promoting Complex-Cyclosome
  • Biological Evolution*
  • Cell Cycle / genetics
  • Cell Cycle / physiology*
  • Chimera
  • DNA Repair*
  • Fungal Proteins / genetics
  • Fungal Proteins / metabolism
  • Genes, Fungal
  • Ligases / genetics*
  • Ligases / metabolism*
  • Models, Biological
  • Molecular Sequence Data
  • Plasmids
  • Saccharomyces cerevisiae / enzymology
  • Saccharomyces cerevisiae / genetics*
  • Saccharomyces cerevisiae / growth & development
  • Saccharomyces cerevisiae Proteins*
  • Sequence Homology, Nucleic Acid
  • Temperature
  • Ubiquitin-Conjugating Enzymes
  • Ubiquitin-Protein Ligase Complexes*
  • Ubiquitin-Protein Ligases

Substances

  • Fungal Proteins
  • Saccharomyces cerevisiae Proteins
  • CDC34 protein, S cerevisiae
  • RAD6 protein, S cerevisiae
  • Ubiquitin-Conjugating Enzymes
  • Ubiquitin-Protein Ligase Complexes
  • Anaphase-Promoting Complex-Cyclosome
  • Ubiquitin-Protein Ligases
  • Ligases