Detection by site-specific disulfide cross-linking of a conformational change in binding of Escherichia coli pyruvate oxidase to lipid bilayers

Y. Y. Chang, J. E. Cronan

Research output: Contribution to journalArticlepeer-review

Abstract

Escherichia coli pyruvate oxidase, a peripheral membrane homotetrameric flavoprotein, exposes its C-terminal lipid binding site in the presence of substrate pyruvate and co-factor thiamine pyrophosphate Mg2+ and binds tightly to phospholipid bilayers during catalysis. Using site-specific disulfide cross-linking, we demonstrate that disulfide cross-links are formed between C termini of D560C pyruvate oxidase and that the degree of cross- linking is greatly increased by the presence of substrate and co-factors indicating a conformational change that results in juxtaposition of two subunit C termini. The cross-linked oxidase is enzymatically active and remains able to associate with lipid micelles. These results argue strongly that lipid bilayer binding of pyruvate oxidase involves pairing of the C termini of two subunits.

Original languageEnglish (US)
Pages (from-to)7896-7901
Number of pages6
JournalJournal of Biological Chemistry
Volume270
Issue number14
DOIs
StatePublished - 1995

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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