title: TP-2Rho Is a Sensitive Solvatochromic Red-Shifted Probe for Monitoring the Interactions between CDK4 and Cyclin D.
abstract: Understanding the intricate steps of protein kinase regulation requires characterization of protein-protein interactions between the catalytic subunit, its regulatory partners and the substrate. Fluorescent probes are useful tools with which to study such interactions and to gain insight into their affinities and specificities. Solvatochromic probes, which display changes in their fluorescence emission in response to changes in the polarity of the medium, are particularly attractive. Here we describe conjugation of a switchable fluorescent dye, TP-2Rho, to peptide and protein derivatives of cyclin-dependent kinase 4 (CDK4) and its application to characterization of the interactions between the catalytic subunit of this kinase, its regulatory partner cyclin D1 and a peptide substrate. We demonstrate the sensitivity of TP-2Rho in relation to of those other dyes used for monitoring peptide-protein and protein-protein interactions. Moreover, we show that TP-Rho-labelled peptides can be introduced into living cells to probe endogenous CDK4/cyclin D.
authors.affiliation: Institut des Biomolécules Max Mousseron-IBMM-CNRS-UMR 5247, Faculté de Pharmacie, Université Montpellier 1, 15, avenue Charles Flahault, 34093, Montpellier, France.
authors.forename: Morgan
authors.initials: M
authors.lastname: Pellerano
authors.affiliation: Institut Curie, PSL Research University, CNRS, INSERM, UMR9187-U1196, 91405, Orsay, France.|Université Paris Sud, Université Paris-Saclay, CNRS, UMR9187-U1196, 91405, Orsay, France.
authors.forename: Delphine
authors.initials: D
authors.lastname: Naud-Martin
authors.affiliation: Institut des Biomolécules Max Mousseron-IBMM-CNRS-UMR 5247, Faculté de Pharmacie, Université Montpellier 1, 15, avenue Charles Flahault, 34093, Montpellier, France.
authors.forename: Marion
authors.initials: M
authors.lastname: Peyressatre
authors.affiliation: Institut des Biomolécules Max Mousseron-IBMM-CNRS-UMR 5247, Faculté de Pharmacie, Université Montpellier 1, 15, avenue Charles Flahault, 34093, Montpellier, France.
authors.forename: Camille
authors.initials: C
authors.lastname: Prével
authors.affiliation: Institut Curie, PSL Research University, CNRS, INSERM, UMR9187-U1196, 91405, Orsay, France.|Université Paris Sud, Université Paris-Saclay, CNRS, UMR9187-U1196, 91405, Orsay, France.
authors.forename: Marie-Paule
authors.initials: MP
authors.lastname: Teulade-Fichou
authors.affiliation: Institut des Biomolécules Max Mousseron-IBMM-CNRS-UMR 5247, Faculté de Pharmacie, Université Montpellier 1, 15, avenue Charles Flahault, 34093, Montpellier, France. may.morris@univ-montp1.fr.
authors.forename: May
authors.initials: M
authors.lastname: Morris
authors.affiliation: Institut Curie, PSL Research University, CNRS, INSERM, UMR9187-U1196, 91405, Orsay, France. florence.mahuteau@curie.fr.|Université Paris Sud, Université Paris-Saclay, CNRS, UMR9187-U1196, 91405, Orsay, France. florence.mahuteau@curie.fr.
authors.forename: Florence
authors.initials: F
authors.lastname: Mahuteau-Betzer
mesh_terms: D056741:Cyclin D / Q000737:chemistry* / Q000378:metabolism*; D051358:Cyclin-Dependent Kinase 4 / Q000737:chemistry* / Q000378:metabolism*; D005456:Fluorescent Dyes / Q000138:chemical synthesis / Q000737:chemistry*; D006367:HeLa Cells; D006801:Humans; D008301:Maleimides / Q000138:chemical synthesis / Q000737:chemistry*; D008958:Models, Molecular; D015394:Molecular Structure; D011485:Protein Binding; D053778:Thiazolidines / Q000138:chemical synthesis / Q000737:chemistry*
chemical_list: D056741:Cyclin D; D005456:Fluorescent Dyes; D008301:Maleimides; C000613454:TP-2Rho-maleimide; D053778:Thiazolidines; C495900:CDK4 protein, human; D051358:Cyclin-Dependent Kinase 4
keywords: CDK4/cyclin D kinase; biosensors; peptide-peptide/protein interactions; protein-protein interactions; solvatochromic fluorescent probes