Structural studies of the acidic transactivation domain of the Vmw65 protein of herpes simplex virus using 1H NMR.

We have overproduced and purified the carboxy-terminal transactivation domain of Vmw65 (VP16) of herpes simplex virus, and studied potential folding of the domain by 1H NMR. Two species of the acidic domain were obtained from the bacterial expression system, and we demonstrate that one of these represents read-through of the natural amber termination codon of the Vmw65 reading frame producing a larger polypeptide. Additional residues in the read-through product were identified by total amino acid analysis and by NMR. Study of the correctly terminated product by 1D NMR gave resonances which were clustered into groups around their random-coil chemical shift positions, and 2D NMR demonstrated that, even in mixed solvents containing up to 80% MeOH, there was very little evidence of secondary structure. Together these results indicate that the isolated acid domain has little if any alpha-helical content of any stable nature. We discuss these results with reference to the demonstrated activity of the acidic domain in a wide variety of polypeptide contexts.