SEQUENCE ANALYSIS OF THE BACTERIAL FLAGELLAR MOTOR

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The proposal is directed to an understanding of biological processes of energy transduction, switching, structure, and assembly, as exemplified in the bacterial flagellar motor. Wild- type and mutant alleles of selected genes coding for components of the flagellar motor of the bacterium Salmonella typhimurium will be sequenced with a view to determining which regions of the proteins, and which amino acids within those regions, are most critical for various aspects of assembly and function; the proposal builds on a large existing body of detailed classical genetic information. There are three main topics: (i) analysis of the five genes (flaAII.2, flaQ, flaN, motA, and motB) that are known to be involved in energy transduction (conversion of proton potential into mechanical work of rotation) and switching (between counterclockwise and clockwise rotation); (ii) comparison of the structural genes for flagellin, hook protein, and hook-associated proteins in an attempt to discover the basis for their recognition by a flagellar-specific export pathway rather than by the conventional N-terminal signal peptide dependent pathway; (iii) analysis of mutant alleles in the flagellin gene that give rise to failure of assembly or to assembly into abnormal helical polymorphs, with a view to identifying residues that are important in the quasi-equivalent inter-subunit interactions that generate the helicity of the filament. As hypotheses emerge from the mutant sequence analysis, they will be tested by generating site- directed mutations.