Y. Pannekoek et al., CONSTRUCTION OF RECOMBINANT NEISSERIAL HSP60 PROTEINS AND MAPPING OF ANTIGENIC DOMAINS, Molecular microbiology, 15(2), 1995, pp. 277-285
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.