SUNY at Albany
June 19-23, 2001
Structure of a Flp recombinase Ð DNA complex
Flp is a very distant member of the lambda integrase family of tyrosine-based site-specific recombinases. Flp catalyzes the inversion of a segment of DNA in the yeast 2micron plasmid, a process that permits the amplification of plasmid copy number. The reaction involves a covalent protein-DNA intermediate and the sequential exchange of pairs of strands.
Using suicide substrates based on mechanistic studies from several laboratories, we have crystallized an intermediate of this reaction. The asymmetric unit of the crystals contains a tetrameric complex, with pseudo fourfold symmetry, and the Holliday junction intermediate is in a nearly square planar conformation. The nearly square planar geometry of the Holliday junction and the fold of the C-terminal catalytic domain are similar to that of the Cre-lox complex determined by the Van Duyne group, but other important features of the complexes are different. Among these are the protein-protein contacts that mediate communication among monomers and enforce the half-of-the-sites activity of the tetramer, and the connectivity of the helix bearing the active site tyrosine, which is domain-swapped in Flp but not in Cre.
Yu Chen(1), Umadevi Narendra(1), Lisa Iype(2), Michael Cox(2), and Phoebe Rice(1)
The University of Chicago(1) and University of Wisconsin(2), Madison
Department of Biochemistry & Molecular Biology; 920 E. 58th St.; Chicago, IL 60637