Characterization of the recD gene of Neisseria gonorrhoeae MS11 and the Effect of recD Inactivation on Pilin Variation and DNA Transformation

摘要

Pilin antigenic variation in Neisseria gonorrhoeae may result followingintrachromosomal recombination between homologous pi/ genes. Despiteextensive study, recA is the only previously characterized gene known to beinvolved in this process. In this study, the gonococcal recD gene, encoding onesubunit of the putative RecBCD holoenzyme, was characterized and its role inpilin variation assessed. The complete recD gene of N. gonorrhoeae M S l l wascloned and its nucleotide sequence determined. The gonococcal recD genecomplemented a defined Escherichia coli recD mutant, based on plaqueformation of bacteriophage 1 and the restoration of ATP-dependent nucleaseactivity. Inactivation of the gonococcal recD gene had no measurable effect oncell viability or survival following UV exposure, but did decrease the frequencyof DNA transformation approximately threefold. The frequency at which nonparentalpilin phenotypes were spawned was 12-fold greater in M S l l recDmutants compared with the parental MS11 re& strain. Similar results wereobtained using recD mutants that were not competent for DNAtransformation. Complementation of the M S l l recD mutant with a wild-typerecD gene copy restored the frequency of pilin phenotypic variation toapproximately wild-type levels. The nucleotide changes at pi/€ in the recDmutants were confined to the variable regions of the gene and were similar tochanges previously attributed to gene conversion.