Research will study the nucleoprotein complexes involved in Tn916 transposition. Tn916 transposable elements are responsible for the conjugative transfer of antibiotic resistance genes in human pathogens, and transpose using a mechanism that is homologous to a wide variety of prokaryotic and eukaryotic site-specific recombination reactions. The nucleic acid chemistry of transposition is mediated by a single transposon encoded protein, integrase. Remarkably, integrase covalently joins two DNA sites separated by greater than 18 kilobases. A key step in this process is the assembly of a recombination complex in which the ends of the transposon are juxtaposed and cleaved by a tetramer of the integrase protein. Assembly of the recombination complex is believed to require multiple integrase-integrase and integrase-DNA interactions. In order to understand the molecular basis of transposition we will use nuclear magnetic resonance (NMR) spectroscopy and biochemical tools to characterize essential integrase-DNA interactions involved in transposition. We will first determine the three dimensional structure and chemical properties of a single integrase-DNA complex that facilitates the formation of the recombination complex. After successfully identifying the molecular interactions that hold the integrase protein tightly onto the transposon sites, we will determine how these interactions might serve as nucleation points for the cooperative binding of other protomers. Finally, we will visualize, at atomic resolution, interactions between DNA-bound integrase protomers. This will be accomplished by determining the three dimensional structure of a protein-DNA complex consisting of two integrase proteins bound to adjacent DNA binding sites.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM057487-04
Application #
6386885
Study Section
Molecular and Cellular Biophysics Study Section (BBCA)
Program Officer
Chin, Jean
Project Start
1998-06-01
Project End
2003-05-31
Budget Start
2001-06-01
Budget End
2002-05-31
Support Year
4
Fiscal Year
2001
Total Cost
$168,405
Indirect Cost
Name
University of California Los Angeles
Department
Pharmacology
Type
Schools of Medicine
DUNS #
119132785
City
Los Angeles
State
CA
Country
United States
Zip Code
90095