title: Substitution of basic amino acids in the basic region stabilizes DNA binding by E12 homodimers.
abstract: The E2A gene encodes two alternatively spliced products, E12 and E47. The two proteins differ in their basic helix-loop-helix motifs (bHLH), responsible for DNA binding and dimerization. Although both E12 and E47 can bind to DNA as heterodimers with tissue-specific bHLH proteins, E12 binds to DNA poorly as homodimers. An inhibitory domain in E12 has previously been found to prevent E12 homodimers from binding to DNA. By measuring the dissociation rates using filter binding and electrophoretic mobility shift assays, we have shown here that the inhibitory domain interferes with DNA binding by destabilizing the DNA-protein complexes. Furthermore, we have demonstrated that substitution of basic amino acids (not other amino acids) in the DNA-binding domain of E12 can increase the intrinsic DNA-binding activity of E12 and stabilize the binding complexes, thus alleviating the repression from the inhibitory domain. This ability of basic amino acids to stabilize DNA-binding complexes may be of biological significance in the case of myogenic bHLH proteins, which all possess two more basic amino acids in their DNA binding domain than E12. To function as heterodimers with E12, the myogenic bHLH proteins may need stronger DNA binding domains.
authors.affiliation: Department of Cell Biology, New York University Medical Center, NY 10016, USA.
authors.forename: S J
authors.initials: SJ
authors.lastname: Vitola
authors.forename: A
authors.initials: A
authors.lastname: Wang
authors.forename: X H
authors.initials: XH
authors.lastname: Sun
mesh_terms: D017398:Alternative Splicing; D000595:Amino Acid Sequence; D001483:Base Sequence; D001665:Binding Sites; D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D004355:Drug Stability; D018257:Helix-Loop-Helix Motifs; D007700:Kinetics; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D016296:Mutagenesis; D013329:Structure-Activity Relationship; D051820:TCF Transcription Factors; D058125:Transcription Factor 7-Like 1 Protein; D014157:Transcription Factors
chemical_list: D004268:DNA-Binding Proteins; D046911:Macromolecular Substances; D051820:TCF Transcription Factors; D058125:Transcription Factor 7-Like 1 Protein; D014157:Transcription Factors; D004247:DNA