Isolation and characterization of dynein ATPase from bull spermatozoa.

Dialysis of demembranated bull spermatozoa against a low-salt buffer resulted in the solubilization of the outer dynein arms and 15-25% of the total ATPase activity. Low-salt extracts contained three high-Mr peptides with Mr values above 300,000. The ATPase activity was associated with two particles sedimenting at 19 S and 12 S. The heavier particle contained two major high-Mr peptides with Mr values above 300,000, one major and one minor intermediate peptides with Mr values of 91,000 and 140,000 respectively and lower-Mr peptides. The 12 S particle contained one high-Mr peptide positioned between the two heavy peptides of the 19 S particle. Even though the peptide compositions of these two particles were different, the enzymic properties of their ATPases were similar. Both particles hydrolysed in the following preference order: ATP greater than CTP greater than UTP greater than ITP greater than GTP. ATPase activities were not affected by ouabain and oligomycin but were inhibited by vanadate, erythro-9-[3-(2-hydroxynonyl)]adenine and EDTA. Enzyme activities were dependent on the presence of a bivalent cation with the following preference order: Mn2+ greater than Mg2+ greater than Ca2+ greater than Ni2+. Optimal activity was observed between pH 6.5 and 9.5. The Km for ATP ranged from 40 to 50 microM for both 19 S and 12 S ATPases. These results suggest that the 12 S and 19 S particles are dyneins from outer dynein arms.