Surface exposure and species specificity of an immunoreactive domain of a 66-kilodalton outer membrane protein (P66) of the Borrelia spp. that cause Lyme disease

Abstract

A chromosomally encoded 66-kDa protein (P66) of Borrelia spp. that cause Lyme disease has previously been shown to be associated with the spirochetal outer membrane. A topological model of P66 predicts a surface-exposed fragment which links the N- and C-terminal intramembranous domains of the protein (J. Bunikis, L. Noppa, and S. Bergström, FEMS Microbiol. Lett. 131:139-145, 1995). In the present study, an immunogenic determinant of P66 was identified by a comparison of the immunoreactivities of different fragments of P66 generated either by proteolytic treatment of intact spirochetes or as recombinant proteins expressed in Escherichia coli. The immune response to P66 during natural infection was found to be directed against the predicted surface domain which comprises amino acids at positions 454 through 491. A sequence comparison revealed considerable polymorphism of the surface domains of P66 proteins of different Lyme disease-causing Borrelia species. Five sequence patterns of this domain were observed in the B. garinii strains studied. In contrast, sequences of the relevant part of P66 of the B. afzelii and B. burgdorferi sensu stricto isolates studied were identical within the respective species. In immunoblotting, 5 of 17 (29.4%) sera from North American patients with early disseminated or persistent Lyme disease reacted against P66 of B. burgdorferi sensu stricto B31. These sera, however, failed to recognize P66 of B. afzelii and B. garinii, as well as an analog of P66 in the relapsing fever agent, B. hermsii. In conclusion, the topological model of P66 is supported by the demonstration of an apparent surface localization of an immunoreactive domain of this protein. Furthermore, analogous to the plasmid-encoded borrelial outer surface proteins, the predicted surface-exposed portion of chromosomally encoded P66 appears to be antigenically heterogenous.