The resolvase encoded by Xanthomonas campestris transposable element ISXc5 constitutes a new subfamily closely related to DNA invertases.

BACKGROUND

Conservative site-specific recombination is responsible for the resolution of cointegrates which result during the transposition of class II transposable elements. Resolution is catalysed by a transposon-encoded recombinase, resolvase, that belongs to a large family of recombinases, including DNA invertases. Resolvases and the related invertases are likely to employ similar reaction mechanisms during recombination. There are important differences, however. Resolvases require two accessory DNA binding sites within each of the two directly repeated recombination sites. Invertases instead need a host factor, Fis, and an enhancer type DNA sequence, in addition to two inversely orientated recombination sites.

RESULTS

The resolvase encoded by transposable element ISXc5 from the gram-negative phytopathogen Xanthomonas campestris shows two features which distinguish it from other known resolvases. First, it is more closely phylogenetically related to invertases than other resolvases. In particular, two functionally important regions seem highly conserved between this resolvase and members of the invertase subfamily. Second, the enzyme exhibits a large extension of its carboxy-terminal domain with unknown function. We purified ISXc5 resolvase and analysed its resolution reaction in vitro. Our biochemical and DNA topological analysis reveals that critical features of resolution are similar, if not identical, to that carried out by gammadelta resolvase. However, despite its apparent similarity to invertases, we were unable to detect recombination on standard substrates for DNA inversion, in either the presence or absence of Fis.

CONCLUSIONS

ISXc5 resolvase employs a reaction mechanism which is common to members of the resolvase family. Its position near the evolutionary borderline to invertases and its high degree of identity within two functionally important regions with members of the DNA invertase subfamily suggest that only a few replacements of critical residues may suffice to convert this resolvase into a functional, possibly Fis-dependent invertase.