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dc.contributor.authorHarborne, Steven PDen
dc.contributor.authorKing, Martin Sen
dc.contributor.authorKunji, Edmunden
dc.date.accessioned2021-03-25T15:59:31Z
dc.date.available2021-03-25T15:59:31Z
dc.date.issued2020-01en
dc.identifier.issn1064-3745
dc.identifier.urihttps://www.repository.cam.ac.uk/handle/1810/319193
dc.description.abstractThere are very few generic methods to assess the stability and functional properties of membrane proteins solubilized in detergent. For this purpose, a thiol-reactive fluorochrome N-[4-(7-diethylamino-4-methyl-3-coumarinyl)phenyl]maleimide (CPM) can be used. An unfolding profile is obtained when the fluorochrome becomes fluorescent on reaction with cysteine residues that have been exposed during thermal denaturation of the protein population. The method was initially developed to optimize the stability of membrane proteins for crystallization studies, but in the course of our work we found many other applications. First, the assay can be used to study the binding of inhibitors, substrates, lipids, and other effectors to membrane proteins. Second, the assay can be used to understand the dynamics of proteins, allowing states to be defined by changes in accessibility of cysteine residues or by changes in specific amino acid interactions. Finally, the assay can be used to study state-dependent domain interactions, for example, as part of regulatory mechanisms. The CPM thermostability assay represents a broadly applicable and versatile tool for a wide range of applications in the functional and structural analysis of membrane proteins.
dc.format.mediumPrinten
dc.languageengen
dc.rightsAll rights reserved
dc.subjectHumansen
dc.subjectBacteriaen
dc.subjectCoumarinsen
dc.subjectAdenomatous Polyposis Coli Proteinen
dc.subjectMembrane Proteinsen
dc.subjectFluorescent Dyesen
dc.subjectDetergentsen
dc.subjectTemperatureen
dc.subjectProtein Denaturationen
dc.subjectProtein Stabilityen
dc.titleThermostability Assays: a Generic and Versatile Tool for Studying the Functional and Structural Properties of Membrane Proteins in Detergents.en
dc.typeArticle
prism.endingPage121
prism.publicationDate2020en
prism.publicationNameMethods in molecular biology (Clifton, N.J.)en
prism.startingPage105
prism.volume2168en
dc.identifier.doi10.17863/CAM.66313
dcterms.dateAccepted2020-03-16en
rioxxterms.versionofrecord10.1007/978-1-0716-0724-4_5en
rioxxterms.versionAM
rioxxterms.licenseref.urihttp://www.rioxx.net/licenses/all-rights-reserveden
rioxxterms.licenseref.startdate2020-01en
dc.contributor.orcidKunji, Edmund [0000-0002-0610-4500]
dc.identifier.eissn1940-6029
rioxxterms.typeJournal Article/Reviewen
pubs.funder-project-idMRC (MC_UU_00015/1)
rioxxterms.freetoread.startdate2021-01-31


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