Epitope mapping of a monoclonal antibody specific for human herpesvirus 6 variant A immediate-early 2 protein.
BACKGROUND
Human herpesvirus 6 (HHV-6) variants A and B are distinct viruses that differ in their biological properties and association to disease. Diagnostic tools able to discriminate between these two variants and between active or latent HHV-6 infection are much needed. In our effort to develop variant-specific antibodies against HHV-6 immediate-early (IE) proteins, we had previously generated P6H8, a monoclonal antibody (mAb) reacting specifically with the immediate-early 2 (IE2) proteins from HHV-6A.


OBJECTIVE
To characterize the P6H8 HHV-6 variant specific mAb and evaluate its potential as part of a variant-specific diagnostic tool for HHV-6A infection. Consequently, our objective was to map the epitope recognized by P6H8.


METHODS
In order to map P6H8 reactivity by Western blotting, we generated deletion mutants of IE2 protein as well as various GST-IE2 fusion proteins. HHV-6A infected cells were used to demonstrate P6H8 reactivity against native IE2. A synthetic peptide corresponding to the P6H8 epitope was used to confirm our results and block P6H8 reactivity.


RESULTS
We mapped the P6H8 epitope to amino acids 1078-1089 of HHV-6A IE2. A peptide (FTPFYYQSSRTR) recreating this epitope was effective in blocking the recognition of both native and recombinant IE2 by P6H8.


CONCLUSIONS
Our work provides a precise characterization of the P6H8 mAb and its specificity toward the IE2 protein of HHV-6 variant A which could prove useful for the differential diagnostic of active infection by HHV-6.
Tomoiu
Andru
A
Laboratory of Virology, Rheumatology and Immunology Research Center, CHUQ Research Center, Quebec, Que., Canada.
Flamand
Louis
L
D000911:Antibodies, Monoclonal / Q000276:immunology*; D000914:Antibodies, Viral / Q000276:immunology*; D002460:Cell Line; D018604:Epitope Mapping; D015654:Herpesvirus 6, Human / Q000276:immunology*; D006801:Humans; D017874:Immediate-Early Proteins / Q000235:genetics / Q000276:immunology*; D009842:Oligopeptides / Q000138:chemical synthesis / Q000276:immunology; D019349:Roseolovirus Infections / Q000175:diagnosis / Q000821:virology*; D012680:Sensitivity and Specificity; D017384:Sequence Deletion
D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; D017874:Immediate-Early Proteins; D009842:Oligopeptides