Characterization of putative mutagenesis cassettes in Sinorhizobium meliloti (927.1)

The goal of this research is to biochemically characterize chromosomal and plasmid-encoded members of Y-family DNA polymerases and putative mutagenesis cassettes from the nitrogen-fixing plant symbiotic bacterium Sinorhizobium meliloti. Y-family translesion polymerases are recruited to sites of DNA damage to replicate damaged DNA that may cause disruption to the replication fork. ImuC is a C family DNA polymerase with a putative function in mutagenic DNA replication; while ImuA and ImuB appear to be important for ImuC function, their roles have yet to be elucidated. S. meliloti harbors the Y family DNA polymerase dinB and the imuABC mutagenesis cassette encoded on both the chromosome and one of the symbiotic plasmids. All of the genes of the putative mutagenesis cassettes have been subcloned into two different plasmid vectors for in vivo and in vitro experiments. We are currently optimizing conditions for protein expression in E. coli. We are also carrying out assays to determine the ability of these gene...