title: Cyan fluorescent protein carries a constitutive mutation that prevents its dimerization.
abstract: The tendency of GFP-like fluorescent proteins to dimerize in vitro is a permanent concern as it may lead to artifacts in FRET imaging applications. However, we have found recently that CFP and YFP (the couple of GFP variants mostly used in FRET studies) show no trace of association in the cytosol of living cells up to millimolar concentrations. In this study, we investigated the oligomerization properties of purified CFP, by fluorescence anisotropy and sedimentation velocity. Surprisingly, we found that CFP has a much weaker homoaffinity than other fluorescent proteins (K(d) ≥ 3 × 10(-3) M), and that this is due to the constitutive N146I mutation, originally introduced into CFP to improve its brightness.
authors.affiliation: Laboratoire de Chimie Physique, Université Paris-Sud and CNRS (UMR 8000), Orsay, France. agathe.espagne@u-psud.fr
authors.forename: Agathe
authors.initials: A
authors.lastname: Espagne
authors.forename: Marie
authors.initials: M
authors.lastname: Erard
authors.forename: Karine
authors.initials: K
authors.lastname: Madiona
authors.forename: Valérie
authors.initials: V
authors.lastname: Derrien
authors.forename: Gabriella
authors.initials: G
authors.lastname: Jonasson
authors.forename: Bernard
authors.initials: B
authors.lastname: Lévy
authors.forename: Hélène
authors.initials: H
authors.lastname: Pasquier
authors.forename: Ronald
authors.initials: R
authors.lastname: Melki
authors.forename: Fabienne
authors.initials: F
authors.lastname: Mérola
mesh_terms: D019943:Amino Acid Substitution / Q000235:genetics; D019281:Dimerization; D005454:Fluorescence Polarization; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods; D005456:Fluorescent Dyes / Q000737:chemistry*; D014644:Genetic Variation; D049452:Green Fluorescent Proteins / Q000737:chemistry* / Q000235:genetics*; D008958:Models, Molecular; D009154:Mutation; D011485:Protein Binding; D055503:Protein Multimerization / Q000235:genetics*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics; D014461:Ultracentrifugation
chemical_list: C485182:Cyan Fluorescent Protein; D005456:Fluorescent Dyes; D011994:Recombinant Proteins; D049452:Green Fluorescent Proteins