Functional Domains in Dynein B Heavy Chain

The dynein ATPases are complex, multisubunit enzymes that generate the forces for a variety of microtubule-based movements. The function of the different dynein subunits in the generation of motility is largely unknown, but the study of mutants with defective flagellar motility in Chlamydomonas reinhardtii has identified several genes that alter the structure and function of the outer dynein arm. One of these genes, the sup-pf-1 locus, is the structural gene for the B subunit of the outer arm complex. The mechanisms by which the B chain contributes to dynein activity and flagellar motility will be studied by the analysis of wild-type and mutant alleles of the sup-pf-1 locus. The specific aims are: 1) To characterize the microtubule binding and translocation properties of wild-type and mutant B subunits by in vitro assays. 2) To determine the physical locations of the microtubule binding site (s) and the sup-pf-1 regulatory regions by photolytic and proteolytic cleavage techniques. 3) To identify the position of specific functional domains within the DNA sequence of the B chain gene, and to characterize the DNA sequences in both mutant and wild-type alleles. This combined genetic, biochemical, and molecular approach will provide new insights into the basic mechanisms of dynein-based motility.