Biochemistry

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    Protocols in Current Issue
    Proximity Ligation Assay for the Investigation of the Intramolecular Interaction of ELMO1
    Authors:  Wai Wa Ray Chan, Dik Long Dennis Chau, Wen Li and Kwok-Fai Lau, date: 12/05/2019, view: 21, Q&A: 0
    Intramolecular interaction is a common mechanism that regulates protein activities. Conventionally, such interactions are investigated by classical in vitro biochemical assays. Here, we describe a protocol for studying the intramolecular ...
    Detection of in vivo Protein Interactions in All Bacterial Components by Förster Resonance Energy Transfer with the Superfolder mTurquoise2 ox-mNeongreen FRET Pair
    Authors:  Nils Y. Meiresonne, Elisa Consoli, Laureen M.Y. Mertens and Tanneke den Blaauwen, date: 12/05/2019, view: 17, Q&A: 0
    This protocol was developed to qualitatively and quantitatively detect protein-protein interactions in all compartments of Escherichia coli by Förster Resonance Energy Transfer (FRET) using the Superfolder mTurquoise2 ox-mNeonGreen FRET ...
    Intracellular cAMP Measurements in Candida albicans Biofilms
    Authors:  Liuliu Jiang, Shengyan Chen, Kairui Sun, Peng Zhou and Xin Wei, date: 12/05/2019, view: 14, Q&A: 0
    Candida albicans is the most common cause of fungal infections worldwide. Infection by C. albicans is closely associated with its ability to form a biofilm, closely packed communities of cells attached to the surfaces of human ...
    Proximity Ligation Assay for the Investigation of the Intramolecular Interaction of ELMO1
    Authors:  Wai Wa Ray Chan, Dik Long Dennis Chau, Wen Li and Kwok-Fai Lau, date: 12/05/2019, view: 21, Q&A: 0
    [Abstract] Intramolecular interaction is a common mechanism that regulates protein activities. Conventionally, such interactions are investigated by classical in vitro biochemical assays. Here, we describe a protocol for studying the intramolecular interaction of cell motility and engulfment 1 (ELMO1) in mammalian cells by using proximity ligation ...
    Detection of in vivo Protein Interactions in All Bacterial Components by Förster Resonance Energy Transfer with the Superfolder mTurquoise2 ox-mNeongreen FRET Pair
    Authors:  Nils Y. Meiresonne, Elisa Consoli, Laureen M.Y. Mertens and Tanneke den Blaauwen, date: 12/05/2019, view: 17, Q&A: 0
    [Abstract] This protocol was developed to qualitatively and quantitatively detect protein-protein interactions in all compartments of Escherichia coli by Förster Resonance Energy Transfer (FRET) using the Superfolder mTurquoise2 ox-mNeonGreen FRET pair (sfTq2ox-mNG). This FRET pair has more than twice the detection range for FRET ...
    Intracellular cAMP Measurements in Candida albicans Biofilms
    Authors:  Liuliu Jiang, Shengyan Chen, Kairui Sun, Peng Zhou and Xin Wei, date: 12/05/2019, view: 14, Q&A: 0
    [Abstract] Candida albicans is the most common cause of fungal infections worldwide. Infection by C. albicans is closely associated with its ability to form a biofilm, closely packed communities of cells attached to the surfaces of human tissues and implanted devices, in or on the host. When tested for susceptibility to antifungals, such as ...
    SarkoSpin: A Technique for Biochemical Isolation and Characterization of Pathological TDP-43 Aggregates
    Authors:  Manuela Pérez-Berlanga, Florent Laferrière and Magdalini Polymenidou, date: 11/20/2019, view: 492, Q&A: 0
    [Abstract] TDP-43 is the main aggregating protein in neurodegenerative diseases such as amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD). Aggregated TDP-43 is resistant to diverse detergent solubilization, yet physiological TDP-43 and other abundant proteins commonly co-purify with pathological TDP-43. This mixed isolation has ...
    ELISA Based Protein Ubiquitylation Measurement
    Authors:  Yuka Kamada, Ryosuke Fukuda and Tsukasa Okiyoneda, date: 11/20/2019, view: 343, Q&A: 0
    [Abstract] Ubiquitylation is a common post-translational modification of cellular proteins that results in proteasomal and lysosomal degradations. Ubiquitylation is generally measured by methods such as immunoblotting using anti-ubiquitin antibodies after isolating the protein-of-interest by denaturing immunoprecipitation. The following protocol can be used ...
    Imaging VIPER-labeled Cellular Proteins by Correlative Light and Electron Microscopy
    [Abstract] Advances in fluorescence microscopy (FM), electron microscopy (EM), and correlative light and EM (CLEM) offer unprecedented opportunities for studying diverse proteins and nanostructures involved in fundamental cell biology. It is now possible to visualize and quantify the spatial organization of cellular proteins and other macromolecules by FM, ...
    Implementing VIPER for Imaging Cellular Proteins by Fluorescence Microscopy
    Authors:  Julia K. Doh, Caroline A. Enns and Kimberly E. Beatty, date: 11/05/2019, view: 243, Q&A: 0
    [Abstract] Genetically-encoded tags are useful tools for multicolor and multi-scale cellular imaging. Versatile Interacting Peptide (VIP) tags, such as VIPER, are new genetically-encoded tags that can be used in various imaging applications. VIP tags consist of a coiled-coil heterodimer, with one peptide serving as the genetic tag and the other (“probe ...
    Generation of CoilR Probe Peptides for VIPER-labeling of Cellular Proteins
    Authors:  Julia K. Doh, Savannah J. Tobin and Kimberly E. Beatty, date: 11/05/2019, view: 274, Q&A: 0
    [Abstract] Versatile Interacting Peptide (VIP) tags are a new class of genetically-encoded tag designed for imaging cellular proteins by fluorescence and electron microscopy. In 2018, we reported the VIPER tag (Doh et al., 2018), which contains two elements: a genetically-encoded peptide tag (i.e., CoilE) and a probe peptide (i.e., ...
    F-actin Bundle Sedimentation Assay
    Authors:  Shan-Shan Lin, Mei-Chun Chuang and Ya-Wen Liu, date: 11/05/2019, view: 255, Q&A: 0
    [Abstract] Understanding the molecular mechanism governing the higher-order regulation of actin dynamics requires chemically-defined and quantitative assays. Recently, the membrane remodeling large GTPase, dynamin, has been identified as a new actin cross-linking molecule. Dynamin regulates actin cytoskeleton through binding to, self-assembling around, and ...
    Isolation and Imaging of His- and RFP-tagged Amyloid-like Proteins from Caenorhabditis elegans by TEM and SIM
    Authors:  Amberley D. Stephens, Meng Lu and Gabriele S. Kaminski Schierle, date: 11/05/2019, view: 335, Q&A: 0
    [Abstract] In our recently published paper, we highlight that during normal aging of C. elegans age-dependent aggregates of proteins form and lead to functional decline of tissues. The protocol described here details the isolation of two proteins from C. elegans in their aggregated amyloid-like form, casein kinase I isoform alpha (KIN-19) ...



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