The long-range goal of the planned research is the understanding of the roles of helix destablizing proteins (single-strand DNA binding proteins) in fundamental processes such as DNA replication, genetic recombination, DNA repair and bacterial conjugation. More specifically, the functions of a newly-discovered gene product from the F sex factor of Escherichia coli will be investigated. This product, SSF (from the ssf gene), is a single-strand DNA binding protein which can complement a temperature-sensitive defect in the closely related E. coli chromosomal gene product, SSB (from the ssb gene). Using site-specific mutagenesis, chromosomal ssb- mutants will be isolated which are completely defective or conditionally suppressible for SSB function, so that solely SSF function can be determined in vivo for its capacity in bacterial and phage DNA replication, DNA repair, SOS induction, genetic recombination and conjugation. A series of representative drug resistance (R) plasmids will be screened for similar ssb-complementing functions. Putative single-strand DNA binding protein genes will then be cloned from selected R factors. Protein products from these cloned fragments will be analyzed using electrophoresis of maxicell extracts, and elution through denatured DNA-cellulose columns. Comparisons of functional activities and DNA sequences will be made in order to develop an understanding of critical functional domains of this series of proteins. Certain E. coli chromosomal suppressors of ssb- mutations will also be analyzed, genetically and functionally, in a further effort to broaden the understanding of this class of DNA binding proteins and other gene products which could interact with them.
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| Golub, E I; Low, K B (1985) Conjugative plasmids of enteric bacteria from many different incompatibility groups have similar genes for single-stranded DNA-binding proteins. J Bacteriol 162:235-41 |
