Isolation of an inhibitor of actin polymerization from human polymorphonuclear leukocytes.
A large proportion of actin in extracts of human polymorphonuclear leukocytes is unpolymerized in the presence of 0.1 M KCl, a condition expected to promote maximal assembly of purified actin. Raising of the KCl concentration increases the extent of actin polymerization in leukocyte extracts, and, at KCl concentrations greater than or equal to 0.3 M, the polymer concentration is maximal and approaches that of purified muscle actin in 0.1 M KCl solution. After removal of the actin polymers from leukocyte extracts made 0.6 M in KCl, the supernatant fluid contains an activity which inhibits the extent of assembly of purified skeletal muscle actin in 0.1 M KCl solution. We have purified this activity by ion exchange and gel filtration chromatography. The purified inhibitor contains polypeptides of 65,000 and 62,000 daltons in a molar ratio of 1:2 as determined by polyacrylamide gel electrophoresis in dodecyl sulfate. The Stokes radius of 32 A, and S20,w of 4.8 of the native inhibitor suggest that the purified protein is a mixture of monomers of slightly different molecular weight. Substoichiometric concentrations of the inhibitor rapidly prevent the assembly of purified actin in 0.1 m KCl solution in a concentration-dependent manner, and the reduction in final viscosity remains constant over time. Raising the KCl concentration reverses the inhibition, as observed for actin polymerization in unfractionated extracts. The findings are consistent with the conclusion that this inhibitor protein is responsible for the large proportion of unpolymerized actin in extracts of polymorphonuclear leukocytes.
D051339:Actin Depolymerizing Factors; D000199:Actins / Q000378:metabolism*; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D001798:Blood Proteins / Q000302:isolation & purification / Q000494:pharmacology*; D051347:CapZ Actin Capping Protein; D051345:Destrin; D006098:Granulocytes / Q000502:physiology*; D006801:Humans; D007700:Kinetics; D046911:Macromolecular Substances; D008840:Microfilament Proteins; D009132:Muscles / Q000378:metabolism; D009504:Neutrophils / Q000502:physiology*; D011487:Protein Conformation; D011817:Rabbits
D051339:Actin Depolymerizing Factors; D000199:Actins; D000596:Amino Acids; D001798:Blood Proteins; C495842:CAPZA1 protein, human; D051347:CapZ Actin Capping Protein; D051345:Destrin; D046911:Macromolecular Substances; D008840:Microfilament Proteins