IDENTIFICATION AND MAPPING OF THE GENE ENCODING THE GLYCOPROTEIN COMPLEX GP82-GP105 OF HUMAN HERPESVIRUS-6 AND MAPPING OF THE NEUTRALIZING EPITOPE RECOGNIZED BY MONOCLONAL-ANTIBODIES

Monoclonal antibodies (MAbs) 2D4, 2D6, and 13D6 against human herpesvi rus 6 (HHV-6) variant A strain GS recognized virion envelope glycoprot ein complex gp82-gp105 and neutralized the infectivity of HHV-6 varian t A group isolates. A 624-bp genomic fragment (82G) was identified fro m an HHV-6 strain GS genomic library constructed in the lambdagt11 exp ression system by immunoscreening with MAb 2D6. Rabbit antibodies agai nst the fusion protein expressed from the genomic insert recognized gl ycoprotein complex gp82-gp105 from HHV-6-infected cells, thus confirmi ng that the genomic fragment is a portion of the gene(s) that encodes gp82-gp105. This genomic insert hybridized specifically with viral DNA s from HHV-6 variant A strains GS and U1102 under high-stringency cond itions but hybridized with HHV-6 variant B strain Z-29 DNA only under low-stringency conditions. DNA sequence analysis of the insert reveale d a 167-amino-acid single open reading frame with an open 5' end and a stop codon at the 3' end. Hybridization studies with HHV-6A strain U1 102 DNA localized the gp82-gp105-encoding gene to the unique long regi on near the direct repeat at the right end of the genome. To locate th e neutralizing epitope(s) recognized by the MAbs, a series of deletion s from the 3' end of the gene were constructed with exonuclease III, a nd fusion proteins from deletion constructs were tested for reactivity with MAbs in a Western immunoblot assay. Sequencing of deletion const ructs at the reactive-nonreactive transition point localized the epito pe recognized by the three neutralizing MAbs within or near a repeat a mino acid sequence (NIYFNIY) of the putative protein. This repeat sequ ence region is surrounded on either side by two potential N-glycosylat ion sites and three cysteine residues.