Understanding of the mechanism of ciliary and eukaryotic flagellar motility requires detailed additional knowledge about subunit composition, subunit structure, and subunit function if the dynein aarms. New results of outer dynein arm substructure of sea urchin sperm axonemes, as studied by the rapid freeze, deep etch approach, make feasible a description of the subunit organization of the dynein arm. Coupled with rapid freeze capture of reactivated sperm in different beat positions, it will be possible to describe changes in subunit structures associated with bend formation. Methods are proposed which will test the effects of chemical fixation on dynein subunit structure and test the possible artifacts induced by the rapid freeze, deep etch proceduce. Dynein 1 has been selectively extracted from sea urchin sperm axonemes and partially fractionated into subunits. The intact dynein 1 unit appears to be the outer dynein arm. The structural and functional domains of each dynein 1 subunit within the outer arm will be analyzed by antibody mapping studies and by precise structural and chemical rebinding to doublet microtubules. Potentially cyclic binding and release of subunits to microtubules will permit purification for further characterization. The mechanism and control of the dynein arms will be studied by experimental manipulation of the pattern of ATP induced microtubule sliding of axonemes briefly treated with elastase. The focus of the work is on the biology of dynein 1 and its role in ciliary and flagellar motility, but it is anticipated that the results will aid in a general understanding of cytoplasmic microtubule function. Additionally, it is now recognized that certain human respiratory and fertility related ailments are a result of ciliary and flagellar disfunction, due to abnormality of the dynein arms. Fundamental knowledge of dynein function may permit more precise description of the molecular basis of dynein related diseases.
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