title: Genetically Encoded Molecular Tension Probe for Tracing Protein-Protein Interactions in Mammalian Cells.
abstract: Optical imaging of protein-protein interactions (PPIs) facilitates comprehensive elucidation of intracellular molecular events. We demonstrate an optical measure for visualizing molecular tension triggered by any PPI in mammalian cells. Twenty-three kinds of candidate designs were fabricated, in which a full-length artificial luciferase (ALuc) was sandwiched between two model proteins of interest, e.g., FKBP and FRB. One of the designs greatly enhanced the bioluminescence in response to varying concentrations of rapamycin. It is confirmed with negative controls that the elevated bioluminescence is solely motivated from the molecular tension. The probe design was further modified toward eliminating the C-terminal end of ALuc and was found to improve signal-to-background ratios, named "a combinational probe". The utilities were elucidated with detailed substrate selectivity, bioluminescence imaging of live cells, and different PPI models. This study expands capabilities of luciferases as a tool for analyses of molecular dynamics and cell signaling in living subjects.
authors: Research Institute for Environmental Management Technology, National Institute of Advanced Industrial Science and Technology (AIST) , 16-1 Onogawa, Tsukuba 305-8569, Japan.
authors: Sung Bae
authors: SB
authors: Kim
authors: Department of Applied Chemistry, Faculty of Science and Technology, Keio University , 3-14-1 Hiyoshi, Kohoku-ku, Yokohama, Kanagawa 223-8522, Japan.
authors: Ryo
authors: R
authors: Nishihara
authors: Department of Applied Chemistry, Faculty of Science and Technology, Keio University , 3-14-1 Hiyoshi, Kohoku-ku, Yokohama, Kanagawa 223-8522, Japan.
authors: Daniel
authors: D
authors: Citterio
authors: Department of Applied Chemistry, Faculty of Science and Technology, Keio University , 3-14-1 Hiyoshi, Kohoku-ku, Yokohama, Kanagawa 223-8522, Japan.
authors: Koji
authors: K
authors: Suzuki
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001696:Biomechanical Phenomena; D019556:COS Cells; D002522:Chlorocebus aethiops; D006801:Humans; D049410:Luciferases, Renilla / Q000737:chemistry / Q000378:metabolism*; D008163:Luminescent Measurements / Q000379:methods; D015335:Molecular Probes / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D011485:Protein Binding; D025941:Protein Interaction Mapping / Q000379:methods*; D017434:Protein Structure, Tertiary; D048790:Renilla / Q000737:chemistry / Q000201:enzymology; D058570:TOR Serine-Threonine Kinases / Q000737:chemistry / Q000378:metabolism*; D022021:Tacrolimus Binding Proteins / Q000737:chemistry / Q000378:metabolism*
chemical_list: D015335:Molecular Probes; D049410:Luciferases, Renilla; D058570:TOR Serine-Threonine Kinases; D022021:Tacrolimus Binding Proteins