Cofilin induced conformational changes in F-actin expose subdomain 2 to proteolysis

Andras Muhlrad, Dmitry Kudryashov, Y. Michael Peyser, Andrey A. Bobkov, Steve C. Almo, Emil Reisler

Research output: Contribution to journalArticle

43 Scopus citations

Abstract

Cofilin/ADF affects strongly the structure of actin filaments and especially the intermolecular contacts of the DNase I binding loop (D-loop) in subdomain 2. In G-actin, the D-loop is cleaved by subtilisin between Met47 and Gly48, while in F-actin this cleavage is inhibited. Here, we report that yeast cofilin, which is resistant to both subtilisin and trypsin, accelerates greatly the rate of subtilisin cleavage of this loop in F-actin at pH 6.8 and at pH 8.0. Similarly, cofilin accelerates strongly the tryptic cleavage in F-actin of loop 60-69 in subdomain 2, at Arg62 and Lys68. The acceleration of the loops' proteolysis cannot be attributed to an increased treadmilling of F-actin for the following reasons: (i) the rate of subtilisin cleavage is independent of pH between pH 6.8 and 8.0, unlike F-actin depolymerization, which is pH-dependent; (ii) at high concentrations of protease the cleavage rate of F-actin in the presence of cofilin is faster than the rate of monomer dissociation from the pointed end of TRC-labeled F-actin, which limits the rate of treadmilling; and (iii) cofilin also accelerates the rate of subtilisin cleavage of F-actin in which the treadmilling is blocked by interprotomer cross-linking of the D-loop to the C terminus on an adjacent protomer. This suggests a substantial flexibility of the D-loop in the cross-linked F-actin. The increased cleavage rates of the D-loop and loop 60-69 reveal extensive exposure of subdomain 2 in F-actin to proteolytic enzymes by cofilin.

Original languageEnglish (US)
Pages (from-to)1559-1567
Number of pages9
JournalJournal of Molecular Biology
Volume342
Issue number5
DOIs
StatePublished - Oct 1 2004

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Keywords

  • ADF
  • DNase I binding loop
  • actin
  • cofilin
  • protein-protein interaction

ASJC Scopus subject areas

  • Structural Biology
  • Molecular Biology

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