Dynamics of protein-DNA interactions in a Mu transpososome

Abstract

Transposition allows movement of a defined stretch of DNA, a transposon, from one DNA location to another. This process is required for the life-cycles of many viruses, from bacteriophage Mu to HIV; it is spreading antibiotic resistances between bacterial populations; and it is responsible for spontaneous mutations in all the kingdoms of life. Transposition is mediated by a protein, the transposase, encoded by the transposon. DNA sequence signals at the two ends of the transposon activate assembly of a transpososome: a complex that include multiple copies of transposase plus both transposon ends. Transpososome assembly, in turn, activates the DNA cleavage and joining reactions required for transposition. This thesis explores aspects of interactions between one transposase, MuA, and the ends of its transposon DNA, the genome of bacteriophage Mu. The first chapter provides an overview of Mu transposition, with special emphasis on the transpososome. The second chapter shows that in the absence of two proper transposon end sequences, an unrelated sequence can substitute for one of the two ends. This leads to some models about the process of transpososome assembly. The third chapter describes and analyzes 18 specific sequences that substituted for one transposon end. The fourth chapter shows that sequence specificity contributes primarily to the initial stage of transposition, transpososome assembly, rather than to the DNA cleavage and joining steps. The fifth chapter shows that the very last nucleotide of the transposon helps maintain the distinction between the two reactions, DNA cleavage and DNA joining.