title: Studies of receptor tyrosine kinase transmembrane domain interactions: the EmEx-FRET method.
abstract: The energetics of transmembrane (TM) helix dimerization in membranes and the thermodynamic principles behind receptor tyrosine kinase (RTK) TM domain interactions during signal transduction can be studied using Förster resonance energy transfer (FRET). For instance, FRET studies have yielded the stabilities of wild-type fibroblast growth factor receptor 3 (FGFR3) TM domains and two FGFR3 pathogenic mutants, Ala391Glu and Gly380Arg, in the native bilayer environment. To further our understanding of the molecular mechanisms of deregulated FGFR3 signaling underlying different pathologies, we determined the effect of the Gly382Asp FGFR3 mutation, identified in a multiple myeloma cell line, on the energetics of FGFR3 TM domain dimerization. We measured dimerization energetics using a novel FRET acquisition and processing method, termed "emission-excitation FRET (EmEx-FRET)," which improves the precision of thermodynamic measurements of TM helix association. The EmEx-FRET method, verified here by analyzing previously published data for wild-type FGFR3 TM domain, should have broad utility in studies of protein interactions, particularly in cases when the concentrations of fluorophore-tagged molecules cannot be controlled.
authors.affiliation: Department of Materials Science and Engineering, Johns Hopkins University, Baltimore, MD 21218, USA.
authors.forename: Mikhail
authors.initials: M
authors.lastname: Merzlyakov
authors.forename: Lirong
authors.initials: L
authors.lastname: Chen
authors.forename: Kalina
authors.initials: K
authors.lastname: Hristova
mesh_terms: D019943:Amino Acid Substitution; D001665:Binding Sites / Q000235:genetics; D019281:Dimerization; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods*; D009154:Mutation; D017434:Protein Structure, Tertiary; D020794:Receptor Protein-Tyrosine Kinases / Q000737:chemistry* / Q000235:genetics; D051498:Receptor, Fibroblast Growth Factor, Type 3 / Q000737:chemistry / Q000235:genetics
chemical_list: D020794:Receptor Protein-Tyrosine Kinases; D051498:Receptor, Fibroblast Growth Factor, Type 3