CHARACTERIZATION OF HUMAN PAPILLOMAVIRUS TYPE-16 E2 PROTEIN AND SUBDOMAINS EXPRESSED IN INSECT CELLS

The E2 open reading frame of human papillomavirus type 16(HPV-16) enco des a DNA-binding protein which modulates papillomavirus transcription and replication. To investigate the biological and biochemical proper ties of the HPV-16 E2 protein, we have constructed recombinant baculov iruses which express the full-length molecule and individual N- and C- terminal domains in Sf21 insect cells. In this system the full-length E2 protein was phosphorylated and targeted to the insect cell nucleus. A 93 amino acid C-terminal fragment encompassing the DNA binding and dimerization functions of E2 was also translocated to the nucleus but was not modified by phosphorylation. The E2 N-terminal protein accumul ated in the insect cell cytoplasm but was not efficiently phosphorylat ed. The formation of heterodimers between full-length and N-terminally truncated E2 species was observed when Sf21 cells were co-infected wi th recombinant viruses and when homodimers were mixed in vitro, sugges ting that the dimer interface is not sufficiently stable to prevent su bunit exchange in vivo. Both homo- and heterodimeric E2 species were a ble to bind specifically and in any combination to tandem E2 binding s ites from the HPV-16 regulatory region. Furthermore, the HPV-16 E2 pro tein bound to DNA exhibited a distinct susceptibility profile to prona se digestion, potentially contrasting with that reported for BPV-1 E2. These observations suggest that significant structural and functional differences may exist between the BPV/HPV E2 proteins and have implic ations for understanding E2-dependent regulation of transcription and replication. (C) 1995 Academic Press, Inc.