CONSTRUCTION OF RECOMBINANT NEISSERIAL HSP60 PROTEINS AND MAPPING OF ANTIGENIC DOMAINS

Citation
Y. Pannekoek et al., CONSTRUCTION OF RECOMBINANT NEISSERIAL HSP60 PROTEINS AND MAPPING OF ANTIGENIC DOMAINS, Molecular microbiology, 15(2), 1995, pp. 277-285
Citations number
46
Categorie Soggetti
Biology,Microbiology
Journal title
ISSN journal
0950382X
Volume
15
Issue
2
Year of publication
1995
Pages
277 - 285
Database
ISI
SICI code
0950-382X(1995)15:2<277:CORNHP>2.0.ZU;2-K
Abstract
Here we report the cloning and expression, in Escherichia coil, of PCR -amplified DNA encoding the 63-kDa stress-inducible protein of Neisser ia gonorrhoeae strains VP1 and PID2, Neisseria meningitidis 2996 and t he commensal Neisseria flavescens. DNA sequence analysis revealed in a ll cases one open reading frame of 541-544 amino acids corresponding t o a protein of approximately 57 000 Da. The various neisserial protein s were >96% identical at the amino acid level and showed extensive hom ology with proteins belonging to the Hsp60 heat-shock-protein family. We constructed defined glutathione S-transferase fusion polypeptides o f the gonococcal Hsp60 homologue to locate antigenic domains on the re combinant protein. Variation in the immunoreactivity of two monoclonal antibodies recognizing a conserved and a neisseria-unique antigenic H sp60 determinant, respectively, could thus be deduced to result from s ingle amino acid substitutions. Analysis of the antibody response in p atients' sera demonstrated reactivity with the same fusion polypeptide s in six out of nine sera, indicating that neisserial Hsp60 is express ed during the natural infection and that distinct domains on the prote in are immunodominant in vivo.