Interaction of profilin with G-actin and poly(L-proline)

Biochemistry. 1994 Jul 19;33(28):8472-8. doi: 10.1021/bi00194a011.


The interaction of bovine spleen profilin with ATP- and ADP-G-actin and poly(L-proline) has been studied by spectrofluorimetry, analytical ultracentrifugation, and rapid kinetics in low ionic strength buffer. Profilin binding to G-actin is accompanied by a large quenching of tryptophan fluorescence, allowing the measurement of an equilibrium dissociation constant of 0.1-0.2 microM for the 1:1 profilin-actin complex, in which metal ion and nucleotide are bound. Fluorescence quenching monitored the bimolecular reaction between G-actin and profilin, from which association and dissociation rate constants of 45 microM-1 s-1 and 10 s-1 at 20 degrees C could be derived. The tryptophan(s) which are quenched in the profilin-actin complex are no longer accessible to solvent, which points to W356 in actin as a likely candidate, consistent with the 3D structure of the crystalline profilin-actin complex [Schutt, C. E., Myslik, J. C., Rozycki, M. D., Goonesekere, N. C. W., & Lindberg, U. (1993) Nature 365, 810-816]. Upon binding poly(L-proline), the fluorescence of both tyrosines and tryptophans of profilin is enhanced 2.2-fold. A minimum of 10 prolines [three turns of poly(L-proline) helix II] is necessary to obtain binding (KD = 50 microM), the optimum size being larger than 10. Binding of poly(L-proline) is extremely fast, with k+ > 200 microM-1 s-1 at 10 degrees C.(ABSTRACT TRUNCATED AT 250 WORDS)

MeSH terms

  • Actins / metabolism*
  • Adenosine Diphosphate / metabolism
  • Adenosine Triphosphate / metabolism
  • Animals
  • Calcium / metabolism
  • Cations, Divalent
  • Cattle
  • Contractile Proteins*
  • Kinetics
  • Microfilament Proteins / chemistry
  • Microfilament Proteins / metabolism*
  • Osmolar Concentration
  • Peptides / chemistry
  • Peptides / metabolism*
  • Profilins
  • Rabbits
  • Spectrometry, Fluorescence
  • Spleen / chemistry
  • Structure-Activity Relationship
  • Tryptophan / chemistry
  • Ultracentrifugation


  • Actins
  • Cations, Divalent
  • Contractile Proteins
  • Microfilament Proteins
  • Peptides
  • Profilins
  • polyproline
  • Adenosine Diphosphate
  • Tryptophan
  • Adenosine Triphosphate
  • Calcium