Procaryotic expression of phosphorylated tegument protein pp65 of human cytomegalovirus and application of recombinant peptides for immunoblot analyses.
The tegument of human cytomegalovirus (HCMV) contains a phosphorylated protein of 65 kilodaltons, termed pp65, which was reported to carry significant epitopes for the stimulation of the humoral immune response during natural infection. A monoclonal antibody directed against this protein was used to screen a lambda gt11 cDNA library for recombinant polypeptides. Two DNA fragments from purified lambda clones and one fragment from genomic DNA were used for cloning in a bacterial high-level expression vector. The resulting fusion proteins were tested for their reactivity with a panel of monoclonal antibodies directed against pp65 and with polyspecific anti-HCMV rabbit antisera. The binding site for all the monoclonal antibodies tested was found to be contained in one of the recombinant proteins with a viral portion of 26 amino acids. Immunoblot analyses with HCMV-positive human sera revealed that pp65 alone is not a reliable antigen for serodiagnosis but may be very useful in combination with other HCMV proteins.
Plachter
B
B
Institut für Klinische und Molekulare Virologie, Universität Erlangen-Nürnberg, Federal Republic of Germany.
Klages
S
S
Hagelmann
S
S
Britt
W
W
Landini
M P
MP
Jahn
G
G
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D000914:Antibodies, Viral / Q000096:biosynthesis; D015153:Blotting, Western; D002478:Cells, Cultured; D003587:Cytomegalovirus / Q000235:genetics*; D004274:DNA, Recombinant / Q000032:analysis; D015870:Gene Expression; D006801:Humans; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism*; D011817:Rabbits; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D015183:Restriction Mapping; D014759:Viral Envelope Proteins / Q000235:genetics* / Q000276:immunology; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*
D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; D004274:DNA, Recombinant; D010750:Phosphoproteins; D011994:Recombinant Proteins; D014759:Viral Envelope Proteins; D014764:Viral Proteins