IDENTIFICATION OF SINGLE AMINO-ACIDS IN THE HUMAN-PAPILLOMAVIRUS-11 E2 PROTEIN CRITICAL FOR THE TRANSACTIVATION OR REPLICATION FUNCTIONS

The papillomavirus E2 protein is required for viral transcriptional re gulation and replication. The E2 protein has a modular structure with two highly conserved domains, a sequence-specific DNA-binding and dime rization domain and a conserved N-terminus which is important for tran scriptional transactivation, replication, and interaction with the E1 protein to determine which specific amino acids or regions in the N-te rminus were important for the replication or transactivation functions . Single amino acid substitutions were created at highly conserved, hi ghly charged amino acids in the HPV 11 E2 N-terminus. Each amino acid was mutated to a nonpolar alanine residue or a similarly charged amino acid. The mutated E2 proteins were analyzed for their abilities to su pport transcriptional transactivation and transient DNA replication an d to enhance binding of E1 to the origin of replication. Single amino acid substitutions were identified which were defective for either the replication or transactivation functions, which demonstrated that the replication and transactivation functions within the N-terminus are s eparable. In several cases different amino acid substitutions at the s ame site had variable effects on transcription or replication, highlig hting the importance of hydrophobic interactions or side chain structu re at each site. The replication function appeared to correlate with t he ability of E2 to enhance binding of E1 to the origin of replication though these studies also suggest that other functions performed by t he E2 protein may be important for replication. (C) 1998 Academic Pres s.