Identification of the structural and functional domains of MutY, an Escherichia coli DNA mismatch repair enzyme

Raymond C. Manuel, Edmund W. Czerwinski, R. Stephen Lloyd

Research output: Contribution to journalArticle

70 Scopus citations

Abstract

The linear amino acid sequences of the Escherichia coli DNA repair proteins, MutY and endonuclease III, show significant homology, even though these enzymes recognize entirely different substrates. In this study, proteolysis and molecular modeling of MutY were used to elucidate its domain organization. Proteolysis by trypsin cleaved the enzyme into 26- and 13-kDa fragments. NH2-terminal sequencing showed that the p13 domain begins at Gln226, indicating that the COOH-terminal portion of MutY, absent in endonuclease III, is organized as a separate domain. The large p26 domain is almost equivalent to the size of endonuclease III. Binding activity of the p26 domain to a DNA substrate containing an A·G mismatch was comparable with that of the intact enzyme. In vitro studies show that the p26 domain retains adenine glycosylase and AP lyase activity on DNA containing undamaged adenine opposite guanine or 8-oxo-7,8-dihydro-2'-deoxyguanine. Although the activity was somewhat reduced, the above results show that the critical amino acid residues involved in substrate binding and catalysis are present in this domain. The structure predicted by molecular modeling indicates that the region of MutY (Met1-Trp216), which is homologous to endonuclease III exhibits a two domain structure, even though this portion is resistant to proteolysis by trypsin.

Original languageEnglish (US)
Pages (from-to)16218-16226
Number of pages9
JournalJournal of Biological Chemistry
Volume271
Issue number27
DOIs
StatePublished - Jul 29 1996

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Fingerprint Dive into the research topics of 'Identification of the structural and functional domains of MutY, an Escherichia coli DNA mismatch repair enzyme'. Together they form a unique fingerprint.

  • Cite this