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A genetic and physical study of the interdomain linker of E-Coli AraC protein-a trans-subunit communication pathway

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dc.contributor.author Malaga, Fabiana
dc.contributor.author Mayberry, Ory
dc.contributor.author Park, David J.
dc.contributor.author Rodgers, Michael E.
dc.contributor.author Toptygin, Dmitri
dc.contributor.author Schleif, Robert F.
dc.date.accessioned 2019-02-22T14:54:33Z
dc.date.available 2019-02-22T14:54:33Z
dc.date.issued 2016
dc.identifier.uri https://hdl.handle.net/20.500.12866/5679
dc.description.abstract Genetic experiments with full length AraC and biophysical experiments with its dimerization domain plus linker suggest that arabinose binding to the dimerization domain changes the properties of the inter-domain linker which connects the dimerization domain to the DNA binding domain via interactions that do not depend on the DNA binding domain. Normal AraC function was found to tolerate considerable linker sequence alteration excepting proline substitutions. The proline substitutions partially activate transcription even in the absence of arabinose and hint that a structural shift between helix and coil may be involved. To permit fluorescence anisotropy measurements that could detect arabinose-dependent dynamic differences in the linkers, IAEDANS was conjugated to a cysteine residue substituted at the end of the linker of dimerization domain. Arabinose, but not other sugars, decreased the steady-state anisotropy, indicating either an increase in mobility and/or an increase in the fluorescence lifetime of the IAEDANS. Time-resolved fluorescence measurements showed that the arabinose-induced anisotropy decrease did not result from an increase in the excited-state lifetime. Hence arabinose-induced decreases in anisotropy appear to result from increased tumbling of the fluorophore. Arabinose did not decrease the anisotropy in mutants incapable of binding arabinose nor did it alter the anisotropy when IAEDANS was conjugated elsewhere in the dimerization domain. Experiments with heterodimers of the dimerization domain showed that the binding of arabinose to one subunit of the dimer decreases the fluorescence anisotropy of only a fluorophore on the linker of the other subunit. en_US
dc.language.iso eng
dc.publisher Wiley
dc.relation.ispartof urn:issn:1097-0134
dc.rights info:eu-repo/semantics/restrictedAccess
dc.rights.uri https://creativecommons.org/licenses/by-nc-nd/4.0/deed.es
dc.subject AraC protein en_US
dc.subject domain en_US
dc.subject fluorescence anisotropy en_US
dc.subject interdomain linker en_US
dc.title A genetic and physical study of the interdomain linker of E-Coli AraC protein-a trans-subunit communication pathway en_US
dc.type info:eu-repo/semantics/article
dc.identifier.doi https://doi.org/10.1002/prot.24990
dc.subject.ocde https://purl.org/pe-repo/ocde/ford#3.02.00 es_PE
dc.subject.ocde https://purl.org/pe-repo/ocde/ford#1.06.03

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