The organization of the outside end of transposon Tn5

Abstract

The end sequences of the IS50 insertion sequence are known as the outside end (OE) and inside end. These complex ends are related but nonidentical 19-bp sequences that serve as substrates for the activity of the Tn5 transposase. Besides providing the binding site of the transposase, the end sequences of a transposon contain additional types of information necessary for transposition. These additional properties include but are not limited to host protein interaction sites and sites that program synapsis and cleavage events. In order to delineate the properties of the IS50 ends,the base pairs involved in the transposase binding site have been defined. This has been approached through performing a variety of in vitro analyses: a ++hydroxyl radical missing-nucleoside interference experiment, a dimethyl sulfate interference experiment, and an examination of the relative binding affinities of single-site end substitutions. These approaches have led to the conclusion that the transposase binds to two nonsymmetrical regions of the OE, including positions 6 to 9 and 13 to 19. Proper binding occurs along one face of the helix, over two major and minor grooves, and appears to result in a significant bending of the DNA centered approximately 3 bp from the donor DNA-OE junction.