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dc.contributor.authorChapman-Smith, A.-
dc.contributor.authorLutwyche, J.-
dc.contributor.authorWhitelaw, M.-
dc.identifier.citationJournal of Biological Chemistry, 2004; 279(7):5353-5362-
dc.description© 2004 by The American Society for Biochemistry and Molecular Biology-
dc.description.abstractThe basic helix-loop-helix (bHLH) PAS transcriptional regulators control critical developmental and metabolic processes, including transcriptional responses to stimuli such as hypoxia and environmental pollutants, mediated respectively by hypoxia inducible factors (HIF-α) and the dioxin (aryl hydrocarbon) receptor (DR). The bHLH proteins contain a basic DNA binding sequence adjacent to a helix-loop-helix dimerization domain. Dimerization among bHLH.PAS proteins is additionally regulated by the PAS region, which controls the specificity of partner choice such that HIF-α and DR must dimerize with the aryl hydrocarbon nuclear translocator (Arnt) to form functional DNA binding complexes. Here, we have analyzed purified bacterially expressed proteins encompassing the N-terminal bHLH and bHLH.PAS regions of Arnt, DR, and HIF-1α and evaluated the contribution of the PAS domains to DNA binding in vitro. Recovery of functional DNA binding proteins from bacteria was dramatically enhanced by coexpression of the bHLH.PAS regions of DR or HIF-1α with the corresponding region of Arnt. Formation of stable protein-DNA complexes by DR/Arnt and HIF-1α /Arnt heterodimers with their cognate DNA sequences required the PAS A domains and exhibited KD values of 0.4 nM and ~50 nM, respectively. In contrast, the presence of the PAS domains of Arnt had little effect on DNA binding by Arnt homodimers, and these bound DNA with a KD of 45 nM. In the case of the DR, both high affinity DNA binding and dimer stability were specific to its native PAS domain, since a chimera in which the PAS A domain was substituted with the equivalent domain of Arnt generated a destabilized protein that bound DNA poorly.-
dc.description.statementofresponsibilityAnne Chapman-Smith, Jodi K. Lutwyche, and Murray L. Whitelaw-
dc.publisherAmer Soc Biochemistry Molecular Biology Inc-
dc.subjectEscherichia coli-
dc.subjectReceptors, Aryl Hydrocarbon-
dc.subjectTranscription Factors-
dc.subjectEnvironmental Pollutants-
dc.subjectElectrophoresis, Polyacrylamide Gel-
dc.subjectTranscription, Genetic-
dc.subjectProtein Structure, Tertiary-
dc.subjectProtein Binding-
dc.subjectActive Transport, Cell Nucleus-
dc.subjectDose-Response Relationship, Drug-
dc.subjectModels, Genetic-
dc.subjectTime Factors-
dc.subjectHypoxia-Inducible Factor 1, alpha Subunit-
dc.titleContribution of the Per/Arnt/Sim (PAS) domains to DNA binding by the basic helix-loop-helix PAS transcriptional regulators-
dc.typeJournal article-
dc.contributor.organisationCentre for the Molecular Genetics of Development-
Appears in Collections:Aurora harvest 2
Centre for the Molecular Genetics of Development publications
Molecular and Biomedical Science publications

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