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. DNA rearrangement/recombination enzymes alter covalent structure of DNA, by cleaving and rejoining DNA strands. These proteins play important roles in diverse biological contexts, including viral integration into host's genome and maintenance of chromosome integrity. Using x-ray crystallography, we are studying two distinct classes of DNA rearrangement enzymes relevant to human diseases. The first class is the retroviral integrase. Retroviruses, including HIV-1 that causes AIDS, have an RNA genome that is reverse-transcribed into a linear viral DNA upon entering the host cell. Integration of this viral DNA into host's chromosome is an essential step in the lifecycle of retroviruses, and is carried out by the virally encoded integrase (IN) protein. Despite high medical relevance of retroviral IN, no structural information is available for the intact 3-domain IN protein responsible for the concerted integration of two viral DNA ends. We are using crystal structures of the functional 3-domain IN protein critically needed for a better mechanistic understanding of IN-catalyzed reactions. Using Rous Sarcoma Virus (RSV) as a model system, we have obtained a diffraction-quality crystal of 3-domain IN. Beam time is requested for higher resolution data collection and phasing experiments. The second class is the DNA resolvase involved in the maintenance of bacterial linear chromosomes. Some bacterial pathogens, including the Lyme disease spirochete Borrelia burgdorferi, have linear chromosomes with covalently closed hairpin termini. Replication of such linear chromosomes requires resolution of a catenated circular intermediate into unit-length chromosomes, which is carried out by the hairpin-forming DNA resolvase enzyme. Using Borrelia and Agrobacterium systems, we are determining crystal structures of the resolvase-DNA complexes to better understand the mechanism of DNA strand cleavages and subsequent hairpin formation.
Showing the most recent 10 out of 407 publications
