title: Mutational analysis of autonomously functioning trans-activating peptides encoded by adenovirus E1A: evidence for two subdomains.
abstract: We have shown previously that a chemically synthesized adenovirus E1A region 3 peptide of 49 amino acids, protein domain 3 (PD3; residues 140 to 188 of the 289-amino-acid protein), trans activates viral genes in vitro and in vivo. To study structure-function relationships, we synthesized N-terminal deletion and cysteine substitution mutant peptides and tested their activities in a cell microinjection assay. Peptides lacking 1 to 12 N-terminal residues exhibited 5- to 50-fold-reduced molar specific activities, whereas those lacking 16 or 18 residues were inactive. Substitution of each of five PD3 cysteine residues with alanine resulted in substantial losses of activity: mutants in the PD3 N-terminal portion showed 40 to 55% of wild-type activity but required a 20-fold-higher concentration than PD3, whereas those in the C-terminal half were as much less active. These peptide mutant studies suggest the existence of two PD3 functional regions: one, localized in the C-terminal 70 to 75% of the molecule, is essential for trans activation; the other, localized in the N-terminal 25 to 30%, can be overridden to a significant extent at high peptide concentrations.
authors.affiliation: Institute for Molecular Virology, St. Louis University Medical Center, Missouri 63110.
authors.forename: R
authors.initials: R
authors.lastname: Pusztai
authors.forename: P M
authors.initials: PM
authors.lastname: Loewenstein
authors.forename: M
authors.initials: M
authors.lastname: Green
mesh_terms: D017456:Adenovirus Early Proteins; D000260:Adenoviruses, Human / Q000235:genetics*; D000595:Amino Acid Sequence; D004252:DNA Mutational Analysis; D015696:Gene Products, tat; D008969:Molecular Sequence Data; D009154:Mutation; D009856:Oncogene Proteins, Viral / Q000235:genetics*; D010455:Peptides / Q000235:genetics*; D014157:Transcription Factors / Q000235:genetics*
chemical_list: D017456:Adenovirus Early Proteins; D015696:Gene Products, tat; D009856:Oncogene Proteins, Viral; D010455:Peptides; D014157:Transcription Factors