DNA strand transfer reactions catalyzed by vaccinia topoisomerase I

Abstract

Vaccinia virus DNA topoisomerase I forms a 3′-phosphoryl intermediate with duplex DNAs containing the conserved binding/cleavage motif 5′CCCTT↓. Covalently bound enzyme is capable of transferring the incised DNA strand to a heterologous DNA acceptor containing a 5′OH terminus. Both intramolecular and intermolecular religation reactions are catalyzed. Intramolecular strand transfer occurs to the noncleaved strand of the DNA duplex and results in formation of a hairpin loop. Intermolecular religation to an exogenous DNA strand is favored over hairpin formation and requires the potential for base pairing between the acceptor and the noncleaved strand of the donor complex. As few as 4 potential base pairs are sufficient to support intermolecular transfer. These results in vitro are consistent with the proposal that vaccinia topoisomerase can catalyze sequence-specific strand transfer during genetic recombination in vivo (Shuman, S. (1991) Proc. Natl. Acad. Sci. U.S.A. 88, 10104-10108.).