Active-site alkylation destabilizes human O6-alkylguanine DNA alkyltransferase

Protein Sci. 2004 Jan;13(1):301-5. doi: 10.1110/ps.03319404.

Abstract

O(6)-alkylguanine-DNA alkyltransferase (AGT) repairs pro-mutagenic O(6)-alkylguanine and O(4)-alkylthymine lesions in DNA. The alkylated form of the protein is not reactivated; instead, it is rapidly ubiquitinated and degraded. Here, we show that alkylation destabilizes the native fold of the protein by 0.5-1.2 kcal/mole and the DNA-binding function by 0.8-1.4 kcal/mole. On this basis, we propose that destabilization of the native conformational ensemble acts as a signal for ubiquitination.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Alkylation
  • Binding Sites
  • Circular Dichroism
  • DNA / metabolism*
  • Humans
  • O(6)-Methylguanine-DNA Methyltransferase / metabolism*
  • Protein Binding
  • Protein Conformation
  • Protein Denaturation
  • Protein Folding
  • Recombinant Proteins / metabolism
  • Ubiquitin / metabolism
  • Urea / pharmacology

Substances

  • Recombinant Proteins
  • Ubiquitin
  • Urea
  • DNA
  • O(6)-Methylguanine-DNA Methyltransferase