Thymosin-beta(4) changes the conformation and dynamics of actin monomers

Biophys J. 2000 May;78(5):2516-27. doi: 10.1016/S0006-3495(00)76797-X.

Abstract

Thymosin-beta(4) (Tbeta(4)) binds actin monomers stoichiometrically and maintains the bulk of the actin monomer pool in metazoan cells. Tbeta(4) binding quenches the fluorescence of N-iodoacetyl-N'-(5-sulfo-1-naphthyl)ethylenediamine (AEDANS) conjugated to Cys(374) of actin monomers. The K(d) of the actin-Tbeta(4) complex depends on the cation and nucleotide bound to actin but is not affected by the AEDANS probe. The different stabilities are determined primarily by the rates of dissociation. At 25 degrees C, the free energy of Tbeta(4) binding MgATP-actin is primarily enthalpic in origin but entropic for CaATP-actin. Binding is coupled to the dissociation of bound water molecules, which is greater for CaATP-actin than MgATP-actin monomers. Proteolysis of MgATP-actin, but not CaATP-actin, at Gly(46) on subdomain 2 is >12 times faster when Tbeta(4) is bound. The C terminus of Tbeta(4) contacts actin near this cleavage site, at His(40). By tritium exchange, Tbeta(4) slows the exchange rate of approximately eight rapidly exchanging amide protons on actin. We conclude that Tbeta(4) changes the conformation and structural dynamics ("breathing") of actin monomers. The conformational change may reflect the unique ability of Tbeta(4) to sequester actin monomers and inhibit nucleotide exchange.

Publication types

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

MeSH terms

  • Actins / chemistry*
  • Actins / metabolism
  • Animals
  • Binding Sites
  • Biophysical Phenomena
  • Biophysics
  • Circular Dichroism
  • Cross-Linking Reagents
  • Fluorescent Dyes
  • Humans
  • In Vitro Techniques
  • Kinetics
  • Macromolecular Substances
  • Models, Molecular
  • Muscle, Skeletal / chemistry
  • Mutagenesis, Site-Directed
  • Naphthalenesulfonates
  • Osmotic Pressure
  • Protein Binding
  • Protein Conformation
  • Rabbits
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Thermodynamics
  • Thymosin / chemistry*
  • Thymosin / genetics
  • Thymosin / metabolism
  • Tritium
  • Viscosity

Substances

  • Actins
  • Cross-Linking Reagents
  • Fluorescent Dyes
  • Macromolecular Substances
  • Naphthalenesulfonates
  • Recombinant Proteins
  • Tritium
  • 1,5-AEDANS
  • thymosin beta(4)
  • Thymosin