We seek to understand at a molecular level the various ways by which an organism maintains the integrity of its genome while accommodating the need for diversity. Our research efforts currently focus on two related processes, homologous recombination and DNA mismatch repair. We are characterizing a central step in genetic recombination, DNA branch migration. DNA branch migration involves the step-wise breakage and reformation of hydrogen bonds in Watson-Crick base pairs as one DNA strand is exchanged for another. Branch migration dictates the amount of genetic information transferred from one chromosome to another. We have determined the rate of spontaneous branch migration and have found that the structure of the exchange point, known as the Holliday junction, plays a pivotal role in determining the rate of spontaneous DNA branch migration. Currently, we are designing branch migration substrates that contain a positioned nucleosome. Mismatch repair, exemplified by the E. coli methyl-directed mismatch repair pathway, plays critical roles in maintaining the integrity of a genome. Mismatches can arise through DNA replication errors, homologous recombination and spontaneous DNA damage. Components of the bacterial mismatch repair system encoded by the mutS and mutL genes in E. coli, are highly conserved in both prokayotes and eukaryotes with defects in human genes encoding mismatch repair enzymes being implicated in hereditary colon cancer. We are interested in understanding the molecular mechanism involved in mismatch recognition by the MutS protein. To this end, we have cloned a gene encoding a thermostable MutS protein from Thermus aquaticus. Initial characterization of the T. aquaticus MutS protein demonstrates that it has thermostable DNA mismatch binding and ATPase activities.
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