The major surface components of the human pathogen Neisseria gonorrhoeae change dramatically during the course of gonorrheal disease in male volunteers. Changes involve the expression states of particular genes in conjunction with the genetic content of the genes being expressed. The Opa family of outer-membrane proteins change expression states as a result of a unique translational switching mechanism involving the insertion or deletion of variable numbers of pentameric coding repeat elements (CR) within that segment of the gene coding for the signal-peptide of the Opa pre-protein. The direct, tandem repeat elements have the capacity to form an intramolecular triplex (H-DNA) in response to negative supercoiling torsion. The opa CR region is the first naturally occurring prokaryotic DNA sequence shown to adopt this non B-DNA conformation and is also unique in that the sequence occurs within the coding sequence of the gene. The phase variation frequencies of repeated genetic elements which form H-DNA are significantly lower than elements with the same repetitive nature which are unable to form H-DNA. Transcription of both types of repeat elements results in increased phase variation frequencies, but this effect is less pronounced in H-DNA forming repetitive sequences. Transcription of opa genes is constitutive for the 5' end of the gene but shows a strong termination when the genes are in the """"""""off"""""""" configuration. The termination may be linked to the formation of the H-DNA structure and if so, represents a novel termination mechanism which could allow the cell to conserve nucleotide triphosphates. The enzymes responsible for generation of negative supercoiling (gyrA/B) were cloned and are being purified to determine their effect on the transcription of opa and the relationship between transcription and H-DNA formation. A number of Opa proteins have been constitutively expressed as beta-lactamase-Opa fusions in E. coli and have been shown to segregate to the outer-membrane in a conformation resembling that seen in N. gonorrhoeae. Recombinant Opa+ bacteria mimic the interactions seen between human neutrophils and gonococcal strains expressing the analogous Opa. Recombinant strains also show an ability to adhere to certain eukaryotic cell lines in in vitro binding assays.

National Institute of Health (NIH)
National Institute of Allergy and Infectious Diseases (NIAID)
Intramural Research (Z01)
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