CHARACTERIZATION OF THE DNA-BINDING ACTIVITY OF THE E1 AND E2 PROTEINS AND THE E1 E2 COMPLEX OF HUMAN PAPILLOMAVIRUS TYPE-33/

The E1 and E2 proteins of papillomaviruses are essential for the initi ation of viral DNA replication. We have purified the E2 protein of hum an papillomavirus type 33 (HPV-33) by immunoaffinity chromatography. T he purified E2 protein bound with high affinity to all four consensus binding sites of HPV-33 (K-d congruent to 2 x 10(-10) M). A putative E 2 binding site differing at one position in the second stem of the pal indrome was not bound by E2. The E1 protein of HPV-33 purified by affi nity chromatography using glutathione S-transferase as tag displayed s pecific DNA-binding activity in footprint analyses protecting HPV-33 n ucleotides 7896 to 7909/1 to 18 from DNasel digestion. Hypersensitive sites at position 6 on the sense and position 1 on the antisense stran d were observed in the middle of the protected region. An E1/E2 comple x protected the E1 binding site and E2 binding sites from DNasel diges tion suggesting that both proteins retain DNA-binding activity in the complex.