title: A comparative analysis of resonance energy transfer methods for Alzheimer related protein-protein interactions in living cells.
abstract: Fluorescence resonance energy transfer (FRET) and bioluminescence resonance energy transfer (BRET) are extensively used to analyze protein interactions occurring in living cells. Although these two techniques are broadly applied in cellular biology, comparative analysis of their strengths and limitations is lacking. To this end, we analyzed a small network of proteins involved in the amyloidogenic processing of the Alzheimer β-amyloid precursor using FRET based cytometry, BRET, and fluorescence lifetime imaging microscopy (FLIM). Using all three methods, we were able to detect the interactions of the amyloid precursor protein with APBB1, APBB2, and APP itself. And we found an unreported interacting pair, APP-APH1A. In addition, we show that these four interacting pairs exhibit a strong FRET correlation with the acceptor/donor expression ratios. Overall the FRET based cytometry was the most sensitive and reliable approach to screen for new interacting proteins. Therefore, we applied FRET based cytometry to study competitive binding of two proteins, APBB1 and APBB2, with the same APP target.
authors: Neurodegeneration and Applied Microscopy, Institut Pasteur Korea, Seongnam-si, Gyeonggi-do, 463-400, South Korea.
authors: Jiho
authors: J
authors: Kim
authors: Joohyun
authors: J
authors: Lee
authors: Doyoon
authors: D
authors: Kwon
authors: Honggun
authors: H
authors: Lee
authors: Regis
authors: R
authors: Grailhe
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000737:chemistry* / Q000378:metabolism; D016564:Amyloid beta-Protein Precursor / Q000737:chemistry / Q000378:metabolism; D001665:Binding Sites; D004735:Energy Transfer; D005434:Flow Cytometry / Q000379:methods; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods*; D057809:HEK293 Cells; D006801:Humans; D008163:Luminescent Measurements / Q000379:methods*; D008856:Microscopy, Fluorescence / Q000379:methods; D009419:Nerve Tissue Proteins / Q000737:chemistry* / Q000378:metabolism; D009687:Nuclear Proteins / Q000737:chemistry* / Q000378:metabolism
chemical_list: C091042:APBB1 protein, human; C466684:APBB2 protein, human; C545487:APP protein, human; D048868:Adaptor Proteins, Signal Transducing; D016564:Amyloid beta-Protein Precursor; D009419:Nerve Tissue Proteins; D009687:Nuclear Proteins