Studies of the dimerization and domain structure of gamma delta resolvase

gamma delta Resolvase is a site-specific recombinase (20.5 kDa) that catalyzes the resolution of a negatively supercoiled plasmid to a catenated pair of circular DNA products (Reed, R. R. (1981) Cell 25, 713-719). Cross-linking experiments and size exclusion high pressure liquid chromatography analysis of isolated fragments corresponding to specific proteolytic cleavage indicate that the intact enzyme and the large fragment exist in a monomer-dimer equilibrium (KDdimer = 8.0 microM, intact enzyme; KDdimer = 0.1 microM, large fragment) and that the small fragment, which displays DNA binding specificity, is a monomer. Dimerization is further supported by line width comparisons in one-dimensional NMR spectra and determinations of the correlation time of the protein. The one-dimensional proton NMR spectra spectroscopy spectra indicate that the overall structure of the two isolated fragments is highly similar to the structure present in the domains of the intact enzyme. The rotational correlation time of resolvase, determined from relaxation data obtained from each domain, indicates that the small domain has a limited degree of additional motion with respect to the large domain of the enzyme. The monomer-dimer equilibrium and small domain mobility may assist in the binding of resolvase to palindromic-type sites with variable spacers and in subunit exchange during catalysis.