title: Allostery within a transcription coactivator is predominantly mediated through dissociation rate constants.
abstract: The kinase-inducible domain interacting (KIX) domain of CREB binding protein binds to multiple intrinsically disordered transcription factors in vivo at two distinct sites on its surface. Several reports have been made of allosteric communication between these two sites in this well-characterized model system. In this work, we have performed fluorescence stopped-flow measurements to investigate the kinetics of binding of five KIX binding proteins. We find that they all have similar association and dissociation rate constants for complex formation, despite their wide range of intrinsic helical propensities. Furthermore, by careful arrangement of pseudofirst-order conditions, we have been able to show that both association and dissociation rate constants are decreased when a partner is bound at the alternative site. These decreases suggest that positive allosteric effects are not mediated by structural changes in binding sites but rather, through a more general mechanism, largely mediated through dissociation, which we propose is largely related to changes in the flexibility of the KIX domain itself.
authors.affiliation: Department of Chemistry, University of Cambridge, Cambridge CB2 1EW, United Kingdom sls42@cam.ac.uk.
authors.forename: Sarah L
authors.initials: SL
authors.lastname: Shammas
authors.affiliation: Department of Chemistry, University of Cambridge, Cambridge CB2 1EW, United Kingdom.
authors.forename: Alexandra J
authors.initials: AJ
authors.lastname: Travis
authors.affiliation: Department of Chemistry, University of Cambridge, Cambridge CB2 1EW, United Kingdom.
authors.forename: Jane
authors.initials: J
authors.lastname: Clarke
mesh_terms: D000494:Allosteric Regulation; D005453:Fluorescence; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D015534:Trans-Activators / Q000737:chemistry*
chemical_list: D015534:Trans-Activators
keywords: coupled folding and binding; dynamic allostery; induced fit; intrinsic disorder; intrinsically disordered protein
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