SAXS STUDIES ON REACTION PATHWAY OF DNA POLYMERASE

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Will test the feasibility and optimize the reaction conditions on the SAXS studies of DNA polymerases and their complexes. Earlier experiments were successful, and the conditions led to minimize the aggregation were also optimized. All the protein samples in the experiments are purified by the gel- filtration chromatography in the final step of protein preparation, and the purity was checked by SDS-PAGE via silver stanning method. No aggregation were detected in these fractions by these above biochemical methods. DNA substrates are purified by gel electrophoresis. Only the puriest fractions were pooled for the SAXS measurements. We are interested in the structures of protein-DNA and protein-DNA-dNTP complexes, and the data will be used for the omplementary structural studies by NMR.