STRUCTURAL AND FUNCTIONAL-CHARACTERIZATION OF THE HPV16-E7 PROTEIN EXPRESSED IN BACTERIA

The E7 gene of the human papillomaviruses (HPV) encodes a 98-amino aci d, multifunctional nuclear phosphoprotein with functional and structur al similarities to adenovirus E1A and the papovavirus T antigens. E7 i s a viral oncoprotein, which will cooperate with an activated ras onco gene to transform primary rodent cells, and can cooperate with the HPV E6 protein for the efficient immortalization of primary human keratin ocytes. Due to the compelling epidemiological and experimental associa tion between HPV infection and cervical cancer, we have undertaken a d etailed study of the structure of the HPV16 E7 protein. The E7 protein was expressed in Escherichia coli as a native, unfused polypeptide, a nd soluble protein was purified by conventional chromatographic techni ques. The purified protein was assessed for various biochemical and bi ophysical properties. Purified E7 binds the retinoblastoma protein avi dly and specifically, and it can dissociate the E2F transcription fact or when assayed in vitro. Circular dichroism spectroscopy indicated th at E7 reversibly binds Zn2+ and Cd2+, resulting in a substantial incre ase in the alpha-helical content of the metal-bound E7 consistent with the stabilization of a hydrophobic core in the COOH terminus of the p rotein.