Structural comparison and epitope analysis of outer-membrane protein PIA from strains of Neisseria gonorrhoeae with differing serovar specificities
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1Molecular Microbiology Group, University of Southampton Medical School, Southampton General Hospital, UK.
Abstract:
The sequences of the por genes, encoding outer-membrane protein PI, have been obtained from a number of strains of Neisseria gonorrhoeae that express PIA molecules with differing serovar specificities. The inferred amino acid sequences of the mature proteins each comprise 308 residues and show considerable homology, with the degree of sequence variation between PIA molecules being considerably less than seen previously with PIB, but more evenly distributed throughout the molecule. The positions of sequence variation are largely confined to the regions predicted to form one of eight surface-exposed loops, suggesting a more widespread distribution of potential antigenic diversity. The deduced amino acid sequences were used to synthesize peptides for epitope mapping experiments. Some epitopes responsible for serovar specificity or recognized by bactericidal monoclonal antibodies could be identified on the basis of their reactivity with simple linear peptides, whilst others recognized conformational epitopes. By comparison of sequence differences with mAb reactivity it was possible to identify regions that appear to contribute to such determinants, including separated regions of the molecule which together were required for the formation of the conformational epitopes. All the epitopes identified lie at or close to the apices of the predicted surface-exposed loops 1, 3, 6, or 8, focusing attention on these regions as accessible targets for immune attack.
Insights
Neisseria gonorrhoeae outer-membrane protein PI (PIA) sequences reveal conserved regions and surface-exposed loops, offering insights into antigenic diversity and potential immune targets for gonorrhea vaccine development.
Area of Science:
- Microbiology
- Immunology
- Structural Biology
Background:
- Neisseria gonorrhoeae outer-membrane protein PI (PIA) is crucial for bacterial survival and pathogenesis.
- Understanding PIA's structure and antigenic properties is key to developing effective vaccines against gonorrhea.
Purpose of the Study:
- To determine the amino acid sequences of PIA from various N. gonorrhoeae strains.
- To analyze sequence variation and its correlation with serovar specificity and antibody recognition.
- To identify potential antigenic targets on PIA for immune-based interventions.
Main Methods:
- Gene sequencing to obtain por gene sequences encoding PIA.
- In silico analysis of inferred amino acid sequences for homology and variation.
- Peptide synthesis based on deduced sequences for epitope mapping.
- Monoclonal antibody (mAb) reactivity assays to identify epitopes.
Main Results:
- PIA sequences show high homology, with variations concentrated in surface-exposed loops.
- Sequence variation in PIA is less than in PIB but more evenly distributed.
- Epitopes responsible for serovar specificity and mAb recognition were mapped to specific surface loops (1, 3, 6, 8).
- Both linear and conformational epitopes were identified, with conformational epitopes requiring multiple regions.
Conclusions:
- PIA exhibits significant sequence conservation with localized antigenic diversity in surface loops.
- Specific surface-exposed loops (1, 3, 6, 8) are primary sites for antigenic determinants.
- These findings highlight accessible targets on PIA for immune attack and potential vaccine design.