Altering the stability of the Cdc8 overlap region modulates the ability of this tropomyosin to bind co-operatively to actin and regulate myosin

Daniel A. East, Duncan Sousa, Stephen R. Martin, Thomas A. Edwards, William Lehman, Daniel P. Mulvihill

Research output: Contribution to journalArticle

Abstract

Tm(tropomyosin) is an evolutionarily conserved α-helical coiled-coil protein, dimers of which form end-to-end polymers capable of associating with and stabilizing actin filaments, and regulating myosin function. The fission yeast Schizosaccharomyces pombe possesses a single essential Tm, Cdc8, which can be acetylated on its N-terminal methionine residue to increase its affinity for actin and enhance its ability to regulate myosin function. We have designed and generated a number of novel Cdc8 mutant proteins with N-terminal substitutions to explore how stability of the Cdc8 overlap region affects the regulatory function of this Tm. By correlating the stability of each protein, its propensity to form stable polymers, its ability to associate with actin and to regulatemyosin,we have shownthat the stability of the N-terminal of the Cdc8 α-helix is crucial for Tm function. In addition we have identified a novel Cdc8 mutant with increased N-terminal stability, dimers of which are capable of forming Tm polymers significantly longer than the wild-type protein. This protein had a reduced affinity for actin with respect to wild-type, andwas unable to regulate actomyosin interactions. The results of the present paper are consistent with acetylation providing a mechanism for modulating the formation and stability of Cdc8 polymers within the fission yeast cell. The data also provide evidence for a mechanism in which Tm dimers form end-to-end polymers on the actin filament, consistent with a co-operative model for Tm binding to actin.

Original languageEnglish (US)
Pages (from-to)265-273
Number of pages9
JournalBiochemical Journal
Volume438
Issue number2
DOIs
Publication statusPublished - Sep 1 2011
Externally publishedYes

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Keywords

  • Acetylation
  • Cdc8
  • Coiled-coil
  • Fission yeast
  • Schizosaccharomyces pombe
  • Tropomyosin

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology
  • Molecular Biology

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