The long-term objectives of the research plan are to elucidate the structure-function relationships that control the elongation cycle during protein biosynthesis and to develop new antibiotics to inhibit the growth of pathogenic bacteria. For the proposed grant period, the goals include the X-ray diffraction structural analyses of Escherichia coli elongation factor (EF-) Tu complexed to antibiotics and ribosomal components using X-ray diffraction techniques. The complexes already co-crystallized include EF-Tu-GDP-GE22709 A, EF-Tu-GDP-kirromycin and EF-Tu-GDP-daunorubicin. Whenever applicable, the EF-Tu complex structures will be solved by using the 1.9 A structure of a trypsin-modified EF-Tu-GDP as a search molecule in molecular replacement calculations. If required, phasing information will also be obtained using multiple isomorphous replacement methods. The proposed research is designed to impact upon practical applications in the design of new antibiotics as well as to further fundamental knowledge of ribosomal processes. First, the structural studies of EF-Tu-antibiotic complexes will yield atomic information about various antibiotic binding sites. This information will be correlated with biochemical data to understand the inhibitory effect of the antibiotics upon EF-Tu function. The information will also be used, in conjunction with the already solved EF-Tu-GDP-tetracycline structure, to design or identify antibiotics that are either more potent or circumvent known bacterial resistance mechanisms. Secondly, as explained in the proposal, those EF-Tu-specific new conformational states of EF-Tu relevant to its function on the ribosome. The atomic information gleaned from EF-Tu-antibiotic complex models will be combined with structural data on EF-Tu-ribosomal components to obtain an atomic description of EF-Tu interactions with the ribosome during protein elongation.
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