Transposition of Mu DNA: Joining of Mu to target DNA can be uncoupled from cleavage at the ends of Mu

Abstract

Transposition of Mu involves transfer of the 3′ ends of Mu DNA to the 5′ ends of a staggered cut in the target DNA. We find that cleavage at the 3′ ends of Mu DNA precedes cutting of the target DNA. The resulting nicked species exists as a noncovalent nucleoprotein complex in which the two Mu ends are held together. This cleaved donor complex completes strand transfer when a target DNA, Mu B protein, and ATP are provided. Mu end DNA sequences that have been precisely cut at their 3′ ends by a restriction endonuclease, instead of by Mu A protein and HU, are efficiently transferred to a target DNA upon subsequent incubation with Mu A protein, Mu B protein, and ATP. Cleavage of the Mu ends therefore cannot be energetically coupled with joining these ends to a target DNA. We discuss the DNA strand transfer mechanism in view of these results, and propose a model involving direct transfer of the 5′ ends of the cut target DNA, from their original partners, to the 3′ ends of Mu.