INTERACTION OF THE RECEPTOR-BINDING DOMAINS OF PSEUDOMONAS-AERUGINOSAPILI STRAINS PAK, PAO, KB7 AND P1 TO A CROSS-REACTIVE ANTIBODY AND RECEPTOR ANALOG - IMPLICATIONS FOR SYNTHETIC VACCINE DESIGN

The four synthetic peptide antigens, PAK 128-144, PAO 128-144, KB7 128 -144 and P1 126-148, correspond in amino acid sequence to the C-termin al receptor binding regions of four strains (PAK, PAO, KB7, P1) of Pse udomonas aeruginosa pilin. The NMR solution structures of the trans fo rms of the peptides show conserved beta-turns which have been implicat ed in antibody and receptor recognition. The interactions between thes e peptides and a cross-reactive monoclonal antibody, PAK-13, have been studied using two-dimensional H-1 NMR spectroscopy in order to map th e antigenic determinants recognized by the antibody. Residues for whic h spectral changes were observed upon antibody binding differed from p eptide to peptide but were mostly confined to one or both of the turn regions and to the hydrophobic pockets. Conformational changes in the beta-turns and hydrophobic pockets of these peptides upon antibody bin ding were also monitored by examination of the pattern of nuclear Over hauser effects (NOEs) versus transferred nuclear Overhauser effects (T RNOEs) for the free versus the bound peptides. Although TRNOEs develop ed strongly between side chain resonances in the hydrophobic pockets o f the peptides, no additional backbone TRNOEs were observed in the pre sence of antibody, suggesting no major conformational changes in the s econdary structures of the peptides upon binding. This implies a flexi ble antibody combining site, a feature which is discussed with respect to cross-reactivity, strain specificity, and the design of a syntheti c peptide vaccine effective against a broad spectrum of P. aeruginosa strains. The binding of the PAK peptide to a disaccharide receptor ana log, (beta GalNAc(1-4)beta Gal), was also studied using H-1 NMR in ord er to map the ''adhesintope'' recognized by the receptor. Spectral cha nges observed in the peptide spectrum with the binding of receptor wer e similar to those seen for the binding of antibody, suggesting that t he epitope recognized by the antibody is structurally coincident with the adhesintope recognized by the receptor. The relevancy of this resu lt is discussed with respect to immunogenicity versus pathogenicity, a nd the proper design of a vaccine which could prevent the mutational e scape of the pathogen away from the host's defence systems. (C) 1997 A cademic Press Limited.