Pattern of aromatic and hydrophobic amino acids critical for one of two subdomains of the VP16 transcriptional activator.
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.
Genetics Program, Michigan State University, East Lansing 48824-1319.
J L
JL
Regier
F
F
Shen
S J
SJ
Triezenberg
D000595:Amino Acid Sequence; D004252:DNA Mutational Analysis; D015967:Gene Expression Regulation, Viral; D017405:Herpes Simplex Virus Protein Vmw65 / Q000235:genetics / Q000378:metabolism*; D007930:Leucine; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010649:Phenylalanine; D011487:Protein Conformation; D018139:Simplexvirus / Q000235:genetics / Q000378:metabolism*; D013329:Structure-Activity Relationship; D014158:Transcription, Genetic; D014162:Transfection
D017405:Herpes Simplex Virus Protein Vmw65; D010649:Phenylalanine; D007930:Leucine