Abstract
Tn5 transposition is a complicated process that requires the formation of a highly ordered protein–DNA structure, a synaptic complex, to catalyse the movement of a sequence of DNA (transposon) into a target DNA. Much is known about the structure of the synaptic complex and the positioning of protein–DNA contacts, although many protein–DNA contacts remain largely unstudied. In particular, there is little evidence for the positioning of donor DNA and target DNA. In this communication, we describe the isolation and analysis of mutant transposases that have, for the first time, provided genetic and biochemical evidence for the stage-specific positioning of both donor and target DNAs within the synaptic complex. Furthermore, we have provided evidence that some of the amino acids that contact donor DNA also contact target DNA, and therefore suggest that these amino acids help define a bifunctional DNA binding region responsible for these two transposase–DNA binding events.
Highlights
IntroductionDNA transposition is a process in which a transposase protein (Tnp) mediates the movement of genetic informa-tion (a mobile DNA segment or transposon) within a genome
DNA transposition is a process in which a transposase protein (Tnp) mediates the movement of genetic informa-tion within a genome
Much is known about the structure of the synaptic complex and the positioning of protein–DNA contacts, many protein–DNA contacts remain largely unstudied
Summary
DNA transposition is a process in which a transposase protein (Tnp) mediates the movement of genetic informa-tion (a mobile DNA segment or transposon) within a genome. Transposons are widespread and have been found in every branch of life (Craig et al, 2002). These elements play an important role in increasing genetic diversity in a population over time. These elements may carry determinants (such as antibiotic-resistance genes) that allow bacterial populations to quickly adapt to new environmental conditions (Davies, 1994). Tn5 is an excellent model system for studying mobile DNA elements because of the fact that Tn5 transposition is a relatively simple system requiring only Tn5 Tnp, Tn5 DNA (defined by recognition end sequences, ESs), target DNA, Mg++ and water, and because extensive genetic, biochemical and structural data exist for this system
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