Identification of seroreactive regions of the human papillomavirus type 16 proteins E4, E6, E7 and L1

Abstract

Small fragments of the DNA of human papillomavirus type 16 (HPV-16) were randomly cloned into the bacteriophage fd which expresses the resulting peptides as part of its capsid. Antisera raised against different HPV-16 fusion proteins were used for screening of the phage clones and the reacting peptides were determined by sequencing the inserted HPV-16 DNA fragments of the positive recombinants. Seroreactive regions of the proteins derived from the E4, E6, E7 (two regions) and L1 (three regions) open reading frames could be found by this approach. Of these seven regions, four were defined by at least two overlapping inserts, thus limiting the domains to between 10 and 15 amino acids. In the case of the E4 open reading frame, the same region identified by immunoscreening was also found when synthetic overlapping octapeptides were tested by ELISA with the anti-E4 antiserum. Using an approach to predict 'receptor-like' regions within the respective proteins, five of the seven regions were also identified. From the data on these regions, synthetic peptides were produced and used for the detection of antibodies against HPV-16 proteins in human sera by ELISA.

Detection of antibodies to L1, L2, and E4 proteins of human papillomavirus types 6, 11, and 16 by ELISA using synthetic peptides

“…Antibodies against HPV 16 E4 peptide were detected in 15% of HPV 16 infected patients vs. 5% of the controls and in none of the patients infected with other HPVs. These results are consistent with those of Suchankova et al [19921 who detected antibodies against another epitope on HPV 16 E4 protein (amino acids 51-70) in 32% of cervical intraepithelial neoplasia (CIN) patients and in 16% of matched control women and those of Muller et al [1990] who detected anti-HPV 16 E4 antibodies in 43% of immunosuppressed patients. The detection of antibodies to HPV 16 E4 protein in HPV infected individuals is consistent with the observation that mRNA encoding the E4 open reading frame (ORF) is one of the most abundant of the early region transcripts in HPV 16 genital precancers [Crum et al, 19911. As in other studies [Mann et al, 19901, a poor correlation between antibodies to HPV 16 and the presence of HPV 16 DNA was observed.…”

Serological responses to HPV 16 in cervical dysplasia and neoplasia: Correlation of antibodies to E6 with cervical cancer

“…Muller et al (1992) used a peptide ELISA and found that 11% of patients with HPV-16-positive invasive cervical cancer were positive to HPV-16 E6 peptide (aa 1-23), 10% were positive to E6 peptide (aa 8-37) and 16% positive to any E6 peptide, as compared with 3%, 1% and 4% respectively in their control group. The MS-2 E6 protein lacks 9 amino terminal amino acids, but still retains the B-cell epitope identified by Muller et al (1990Muller et al ( , 1992. The presence of additional epitopes in the MS-2 fusion protein used in this study may account for the higher proportions of cervical-cancer patients with antibodies to E6 detected in this study.…”

“…By contrast, Kochel et al (1991) detected anti-E6 antibodies in only one of 23 cervical-cancer patients. This low incidence could be due to the use in their Western blot assay of a E6 p-gal fusion protein which lacks 30 amino terminal amino acids, the region containing the major B-cell epitopes (Muller et al, 1990(Muller et al, ,1992.…”

Antibody response to a synthetic peptide derived from the human papillomavirus type 6/11 L2 protein in recurrent respiratory papillomatosis: Correlation between southern blot hybridization, polymerase chain reaction, and serology

“…HPV 6 genomes in the benign laryngeal lesions did not have this duplication, suggesting that sequence rearrangements in the URR were necessary to confer malignant potential to "low risk' HPVs [Byrne et al, 1987;DiLorenzo et al, 19921. Immunoassays capable of detecting type-specific anti-HPV antibodies in sera of patients could be helpful in the diagnosis of HPV-associated diseases and would be in theory a straightforward and inexpensive alternative to molecular hybridization. Since viral antigens cannot be prepared in sufficient quantities in tissue cultures, fusion proteins and synthetic peptides have been commonly used for detecting HPV-reactive antibodies [Cason et al, 1989;Dillner et al, 1989;Dillner, 1990;Jochmus-Kudielka et al, 1989;Muller et al, 1990;Suchankova et al, 19901. In this study, we have assessed the correlation between serology, Southern blot hybridization, and PCR in detecting HPV in a series of 25 patients with recurrent respiratory papillomatosis.…”