title: A Protein-Protein Interaction Assay FlimPIA Based on the Functional Complementation of Mutant Firefly Luciferases.
abstract: There is a significant focus on detecting and assaying protein-protein interactions (PPIs) in biology and biotechnology. Protein-fragment complementation assay (PCA) is one of the most widely used methods to detect PPI by splitting the enzyme-coding or fluorescent protein-coding polypeptide, as well as Förster resonance energy transfer (FRET). Here, we describe a novel PPI assay FlimPIA (firefly luminescent intermediate-based protein-protein interaction assay) by a unique approach of splitting the two major catalytic steps (half reactions) of firefly luciferase (FLuc).
authors.affiliation: Department of Chemical Science and Engineering, Graduate School of Engineering, Kobe University, 1-1 Rokkodai, Nada, Kobe, 657-8501, Japan.
authors.forename: Yuki
authors.initials: Y
authors.lastname: Ohmuro-Matsuyama
authors.affiliation: Laboratory for Chemistry and Life Science, Tokyo Institute of Technology, 4259-R1-18 Nagatsuta-cho, Midori-ku, Yokohama, 226-8503, Japan. ueda@res.titech.ac.jp.
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Ueda
mesh_terms: D031541:Fluorescence Resonance Energy Transfer; D015870:Gene Expression; D005822:Genetic Vectors / Q000235:genetics; D049409:Luciferases, Firefly / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D008163:Luminescent Measurements / Q000379:methods; D050505:Mutant Proteins; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs; D025941:Protein Interaction Mapping / Q000379:methods*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism
chemical_list: D050505:Mutant Proteins; D011993:Recombinant Fusion Proteins; D049409:Luciferases, Firefly
keywords: Firefly luciferase; Luciferyl adenylate; Protein–protein interaction; Reaction intermediate; Substrate channeling