The long-term objective of this research proposal is to contribute to the biochemical understanding of the mechanism of homologous recombination. The general approach is to enzymatically characterize the purified proteins known to participate in genetic recombination in E. coli and to determine their enzymatic mechanism when acting individually or as component of a more complex system. These proteins include the recA, SSB, and recBCD proteins of E. coli. The specific approach is to pursue physical-biochemical, enzymatic, and structure-functional characterization of a protein that plays a central role in genetic recombination, the recA protein of E. coli. To assess the biological significance of the many recA protein activities and to determine how they contribute to enzymatic function, the biochemical properties of mutant recA proteins will be examined. Finally, in order to better understand how the activities of the recA protein are coordinated with those of SSB and recBCD enzymes to promote the homologous pairing, in vitro assays that require the activities of these proteins will be examined mechanistically. Understanding the molecular mechanism of genetic recombination should shed light aberrant on biological events such as chromosomal translocations and error-prone repair and should contribute to the fundamental biochemical knowledge required to develop useful gene replacement therapies based on homologous recombination.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
7R01AI018987-11
Application #
3128400
Study Section
Biochemistry Study Section (BIO)
Project Start
1982-04-01
Project End
1995-03-31
Budget Start
1991-09-01
Budget End
1992-03-31
Support Year
11
Fiscal Year
1991
Total Cost
Indirect Cost
Name
University of California Davis
Department
Type
Schools of Medicine
DUNS #
094878337
City
Davis
State
CA
Country
United States
Zip Code
95618
Wu, Yun; Kantake, Noriko; Sugiyama, Tomohiko et al. (2008) Rad51 protein controls Rad52-mediated DNA annealing. J Biol Chem 283:14883-92
Seitz, Erica M; Kowalczykowski, Stephen C (2006) Human Rad51 protein displays enhanced homologous pairing of DNA sequences resembling those at genetically unstable loci. Nucleic Acids Res 34:2847-52
Wu, Yun; Sugiyama, Tomohiko; Kowalczykowski, Stephen C (2006) DNA annealing mediated by Rad52 and Rad59 proteins. J Biol Chem 281:15441-9
Kantake, Noriko; Sugiyama, Tomohiko; Kolodner, Richard D et al. (2003) The recombination-deficient mutant RPA (rfa1-t11) is displaced slowly from single-stranded DNA by Rad51 protein. J Biol Chem 278:23410-7
New, James H; Kowalczykowski, Stephen C (2002) Rad52 protein has a second stimulatory role in DNA strand exchange that complements replication protein-A function. J Biol Chem 277:26171-6
Kantake, Noriko; Madiraju, Murty V V M; Sugiyama, Tomohiko et al. (2002) Escherichia coli RecO protein anneals ssDNA complexed with its cognate ssDNA-binding protein: A common step in genetic recombination. Proc Natl Acad Sci U S A 99:15327-32
Solinger, J A; Lutz, G; Sugiyama, T et al. (2001) Rad54 protein stimulates heteroduplex DNA formation in the synaptic phase of DNA strand exchange via specific interactions with the presynaptic Rad51 nucleoprotein filament. J Mol Biol 307:1207-21
Mazin, A V; Bornarth, C J; Solinger, J A et al. (2000) Rad54 protein is targeted to pairing loci by the Rad51 nucleoprotein filament. Mol Cell 6:583-92
Mazin, A V; Zaitseva, E; Sung, P et al. (2000) Tailed duplex DNA is the preferred substrate for Rad51 protein-mediated homologous pairing. EMBO J 19:1148-56
Kowalczykowski, S C (2000) Initiation of genetic recombination and recombination-dependent replication. Trends Biochem Sci 25:156-65

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