title: Mechanism of the interaction of human platelet profilin with actin.
abstract: We have reexamined the interaction of purified platelet profilin with actin and present evidence that simple sequestration of actin monomers in a 1:1 complex with profilin cannot explain many of the effects of profilin on actin assembly. Three different methods to assess binding of profilin to actin show that the complex with platelet actin has a dissociation constant in the range of 1 to 5 microM. The value for muscle actin is similar. When bound to actin, profilin increases the rate constant for dissociation of ATP from actin by 1,000-fold and also increases the rate of dissociation of Ca2+ bound to actin. Kinetic simulation showed that the profilin exchanges between actin monomers on a subsecond time scale that allows it to catalyze nucleotide exchange. On the other hand, polymerization assays give disparate results that are inconsistent with the binding assays and each other: profilin has different effects on elongation at the two ends of actin filaments; profilin inhibits the elongation of platelet actin much more strongly than muscle actin; and simple formation of 1:1 complexes of actin with profilin cannot account for the strong inhibition of spontaneous polymerization. We suggest that the in vitro effects on actin polymerization may be explained by a complex mechanism that includes weak capping of filament ends and catalytic poisoning of nucleation. Although platelets contain only 1 profilin for every 5-10 actin molecules, these complex reactions may allow substoichiometric profilin to have an important influence on actin assembly. We also confirm the observation of I. Lassing and U. Lindberg (1985. Nature [Lond.] 318:472-474) that polyphosphoinositides inhibit the effects of profilin on actin polymerization, so lipid metabolism must also be taken into account when considering the functions of profilin in a cell.
authors.affiliation: Department of Cell Biology and Anatomy, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205.
authors.forename: P J
authors.initials: PJ
authors.lastname: Goldschmidt-Clermont
authors.forename: L M
authors.initials: LM
authors.lastname: Machesky
authors.forename: S K
authors.initials: SK
authors.lastname: Doberstein
authors.forename: T D
authors.initials: TD
authors.lastname: Pollard
mesh_terms: D000199:Actins / Q000302:isolation & purification / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D001792:Blood Platelets / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism; D003198:Computer Simulation; D003285:Contractile Proteins / Q000097:blood; D004985:Ethenoadenosine Triphosphate / Q000031:analogs & derivatives / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D046911:Macromolecular Substances; D008840:Microfilament Proteins / Q000097:blood* / Q000302:isolation & purification / Q000494:pharmacology; D008970:Molecular Weight; D009132:Muscles / Q000378:metabolism; D018129:Phosphatidylinositol Phosphates; D010716:Phosphatidylinositols / Q000494:pharmacology; D051304:Profilins; D011485:Protein Binding; D011817:Rabbits
chemical_list: D000199:Actins; D003285:Contractile Proteins; D004985:Ethenoadenosine Triphosphate; D046911:Macromolecular Substances; D008840:Microfilament Proteins; C495738:PFN1 protein, human; D018129:Phosphatidylinositol Phosphates; D010716:Phosphatidylinositols; D051304:Profilins; C012758:aza-epsilon-ATP; D000255:Adenosine Triphosphate; D002118:Calcium