Identification of two topologically distinct Mu transpososomes: contribution of cis and trans elements to DNA topology

Abstract

Transposition of bacteriophage Mu takes place within nucleoprotein complexes called transpososomes. Assembly of a transpososome requires transposase binding to multiple sites within the L and R ends of Mu and an internal transpositional enhancer E, present on supercoiled DNA. L, R and E interact such that five negative DNA supercoils are trapped within the transpososome. We have carried out topological studies aimed at understanding the contribution of all of these elements to transpososome assembly. We have found that the transposase has an inherent capacity to interwrap distant DNA sites. Under enhancer-independent reaction conditions, two topologically distinct synapses were identified. These studies have revealed that the enhancer as well as the ends, contribute to the topological selectivity of the Mu synapse. We have identified specific end – enhancer interactions critical for assembly. Based on our topological studies as well as other data, we propose a new model for the structure of the 5-noded 3-site Mu complex.