FUNCTIONAL INTERACTION BETWEEN CHLAMYDOMONAS OUTER ARM DYNEIN SUBUNITS - THE GAMMA-SUBUNIT SUPPRESSES THE ATPASE ACTIVITY OF THE ALPHA-BETADIMER

The alpha beta dimer and the gamma subunit of the Chlamydomonas outer arm dynein were solubilized by treating isolated axonemes with 0.6 M K CI, and purified by sucrose density gradient centrifugation. The axone mes were from an ida1 mutant to eliminate contamination of outer arm s ubunits by inner arm dynein I1, and the axonemes were pre-extracted wi th 0.6 M CH3COOK to remove non-dynein protein that might otherwise con taminate outer arm dynein fractions in the sucrose gradient. In additi on, purer fractions of outer arm dynein subunits were obtained by modi fying the centrifugation conditions to take advantage of the propensit y of the dynein to dissociate under high hydrostatic pressure in the p resence of Mg2+. When sucrose gradient fractions containing the gamma subunit were added to a fraction containing the purified alpha beta di mer under conditions expected to promote reassociation of the subunits to form a trimeric outer arm dynein complex [Takada et al., 1992: J. Biochem. 111:758-762], the total ATPase activity of the mixture was su ppressed to a level lower than that of the original alpha beta dimer f raction. The inhibition paralleled the distribution of gamma subunit i n the sucrose gradient, was saturable, and was maximum at an approxima tely equimolar ratio of the gamma subunit to the alpha beta dimer. The se results indicate that when the gamma subunit interacts with the alp ha beta dimer, the latter's ATPase activity is modulated downward. Pre vious results showed that interaction of the alpha subunit with the be ta subunit suppressed the beta subunit's ATPase activity [Pfister and Witman, 1984: J. Biol. Chem. 259:12072-12080]. Thus, the total ATPase activity of the outer arm dynein is dependent upon communication betwe en all three subunits within the arm. (C) 1997 Wiley-Liss, Inc.