Structural features of the transcriptional activation domain of the
herpes simplex virion
protein VP16 were examined by
oligonucleotide-directed mutagenesis. Extensive mutagenesis at position 442 of the truncated
VP16 activation domain (delta 456), normally occupied by a
phenylalanine residue, demonstrated the importance of an
aromatic amino acid at that position. On the basis of an alignment of the
VP16 sequence surrounding Phe-442 and the sequences of other transcriptional activation domains, we subjected
leucine residues at positions 439 and 444 of
VP16 to mutagenesis. Results from these experiments suggest that bulky hydrophobic residues flanking Phe-442 also contribute significantly to the function of the truncated
VP16 activation domain. Restoration of
amino acids 457-490 to various Phe-442 mutants partially restored activity. Although the pattern of
amino acids surrounding Phe-473 resembles that surrounding Phe-442, mutations of Phe-473 did not dramatically affect activity; in fact, Phe-475 appears more sensitive to mutations than does Phe-473. We infer that the two regions of
VP16 (
amino acids 413-456 and 457-490) possess unique structural features, although neither is likely to be an amphipathic alpha-helix or an "acidic blob." These results, considered with previous in vitro activation and inhibition studies, suggest that the two subdomains of
VP16 affect transcription by different mechanisms.