β Recombinase catalyzes inversion and resolution between two inversely oriented six sites on a supercoiled DNA substrate and only inversion on relaxed or linear substrates

Abstract

The β recombinase, in the presence of a chromatin-associated protein such as Hbsu, catalyzes DNA resolution or DNA inversion on supercoiled substrates containing two directly or inversely oriented six sites. Hbsu stabilizes the formation of the recombination complex (Alonso, J.C., Weise, F., and Rojo, F. (1995) J. Biol. Chem. 270, 2938-2945). In this study we show that resolution by β recombinase strictly requires supercoiled DNA, but inversion does not. On a substrate with two inversely oriented six sites, β recombinase catalyzed both resolution and inversion if the DNA was supercoiled but only inversion if the substrate was relaxed or linear. Hbsu was critical for the formation of synaptic complexes; its concentration relative to that of the supercoiled DNA substrate determined whether resolution or inversion products were preferentially formed. The results suggest that the β recombinase forms unproductive short-lived synaptic complexes between two juxtaposed inversely oriented six sites; the presence of 3 to 13 Hbsu dimers per supercoiled DNA molecule would stabilize a synaptic complex with a relative geometry of the six sites allowing β recombinase preferentially to achieve resolution. Supercoiling probably helps to overcome an energetic barrier, since resolution does not occur in relaxed DNA. The presence of >30 Hbsu dimers per DNA molecule probably favors the formation of a recombination complex with a different geometry since the reaction is directed preferentially toward DNA inversion.