Protocols in Current Issue
    Reconstitution of Membrane-associated Components of a G-protein Signaling Pathway on Membrane-coated Nanoparticles (Lipobeads)
    Authors:  Michael J. Irwin, Xin Wang and Rick H. Cote, date: 01/20/2022, view: 1256, Q&A: 0

    G-protein coupled signaling pathways are organized into multi-protein complexes called signalosomes that are located within and on cellular membranes. We describe the use of silica nanoparticles coated with a unilamellar phospholipid bilayer (lipobeads) to reconstitute the activated photoreceptor G-protein α-subunit (Gtα*) with its

    Cell-free Generation of COPII-coated Procollagen I Carriers
    Authors:  Lin Yuan, Satoshi Baba, Kanika Bajaj and Randy Schekman, date: 11/20/2017, view: 7049, Q&A: 0
    [Abstract] The aim of this protocol is to generate COPII-coated procollagen I (PC1) carriers in a cell-free reaction. The COPII-coated PC1 carriers were reconstituted from donor membrane, cytosol, purified recombinant COPII proteins, and nucleotides. This protocol describes the preparation of donor membrane and cytosol, the assembly of the reaction, and the ...
    Membrane Lipid Screen to Identify Molecular Targets of Biomolecules
    Authors:  John R. Jimah, Paul H. Schlesinger and Niraj H. Tolia, date: 08/05/2017, view: 8210, Q&A: 0
    [Abstract] Proteins that bind to and disrupt cell membranes may target specific phospholipids. Here we describe a protocol to identify the lipid targets of proteins and biomolecules. First, we describe a screen to identify lipids in membranes that are specifically bound by the biomolecule of interest. Second, we describe a method for determining if the ...
    Assay to Measure Interactions between Purified Drp1 and Synthetic Liposomes
    Authors:  Yoshihiro Adachi, Kie Itoh, Miho Iijima and Hiromi Sesaki, date: 05/05/2017, view: 9380, Q&A: 0
    [Abstract] A mitochondrion is a dynamic intracellular organelle that actively divides and fuses to control its size, number and shape in cells. A regulated balance between mitochondrial division and fusion is fundamental to the function, distribution and turnover of mitochondria (Roy et al., 2015). Mitochondrial division is mediated by ...
    Liposome Flotation Assays for Phosphoinositide-protein Interaction
    Authors:  Helene Tronchere and Frederic Boal, date: 03/05/2017, view: 17455, Q&A: 0
    [Abstract] Phosphoinositides are rare membrane lipids involved in the control of the major cellular functions and signaling pathways. They are able to recruit specific effector proteins to the cytosolic face of plasma membrane and organelles to coordinate a vast variety of signaling and trafficking processes, as well to maintain specific identity of the ...
    Examination of the Interaction between a Membrane Active Peptide and Artificial Bilayers by Dual Polarisation Interferometry
    Authors:  Jennifer A.E. Payne, Tzong-Hsien Lee, Marilyn A. Anderson and Marie-Isabel Aguilar, date: 01/05/2017, view: 6861, Q&A: 0
    [Abstract] Examining the interaction of peptides with lipid bilayers to determine binding kinetics is often performed using surface plasmon resonance (SPR). Here we describe the technique of dual polarisation interferometry (DPI) that provides not only information on the kinetics of the peptide binding to the bilayer, but also how the peptide affects the ...
    Determining Efficiency and Selectivity of Lipid Extraction by Perturbing Agents from Model Membranes
    Authors:  Michel Lafleur and Alexandre Therrien, date: 11/20/2016, view: 6962, Q&A: 0
    [Abstract] Several membrane-perturbing agents extract lipids from membranes and, in some cases, this lipid efflux is lipid specific. In order to gain a better description of this phenomenon and to detail the intermolecular interactions that are involved, a method has been developed to characterize the extent and the specificity of membrane-lipid extraction ...
    Preparation of Chloroplast Lipid Membrane and Lipid-protein Interaction Assay
    Authors:  Kumiko Okazaki, Shin-ya Miyagishima and Hajime Wada, date: 02/05/2016, view: 9088, Q&A: 0
    [Abstract] Lipid-Protein interaction assay is a method to search lipids, which are bound with proteins in vitro. Since membranes that are spotted with chloroplast lipids such as monogalactosyldiacylglycerol (MGDG), digalactosyldiacylglycerol (DGDG), and sulfoquinovosyldiacylglycerol (SQDG) are not commercially available, we extracted these lipids ...
    In Gel Detection of Lipase Activity in Crude Plant Extracts (Olea europaea)
    [Abstract] Here, we provide a detailed protocol describing a SDS-PAGE based procedure to assay in gel neutral lipase activity. Total protein extracts are separated by SDS-PAGE and gels are treated with lipase substrate-α-naphthyl palmitate. This long-chain fatty acid ester is hydrolysed by lipases present in the gel. The product resulting from this reaction ...
    Competitive ELISA for Protein-lipopolysaccharide (LPS) Binding
    Authors:  Victoria Martínez-Sernández and Florencio M. Ubeira, date: 11/20/2014, view: 13555, Q&A: 1
    [Abstract] Lipopolysaccharide is the major constituent of the outer membrane of gram-negative bacteria and, once released from the bacterial surface into the bloodstream, is a potent activator of the host immune system, which can lead to septic shock. LPS has a hydrophilic region consisting of a repeating oligosaccharide that is strain-specific (O-antigen) ...

    We use cookies on this site to enhance your user experience. By using our website, you are agreeing to allow the storage of cookies on your computer.