Biochemistry

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    Protocols in Current Issue
    Simple Method to Determine Protein Redox State in Arabidopsis thaliana
    Authors:  Keisuke Yoshida and Toru Hisabori, date: 06/05/2019, view: 414, Q&A: 0
    [Abstract] Thiol-based redox regulation is a posttranslational protein modification that plays a key role in many biological aspects. To understand its regulatory functions, we need a method to directly assess protein redox state in vivo. Here we present a simple procedure to determine protein redox state in a model plant Arabidopsis thaliana ...
    Detection of Disulfides in Protein Extracts of Arabidopsis thaliana Using Monobromobimane (mBB)
    Authors:  Shin‐nosuke Hashida and Maki Kawai-Yamada, date: 03/05/2019, view: 754, Q&A: 0
    [Abstract] Thiol-disulfide exchange is a key posttranslational modification, determining the folding process of intra- and inter-protein structures. Thiols can be detected by colorimetric reagents, which are stoichiometrically reduced by free thiols, and by fluorescent adducts, showing fluorescence only after thioester formation. We adapted a simple ...
    Preparation of Silk Fibroin Nanoparticles and Enzyme-Entrapped Silk Fibroin Nanoparticles
    Author:  Yu-Qing Zhang, date: 12/20/2018, view: 1433, Q&A: 0
    [Abstract] After silk fiber is degummed in boiling 0.2% Na2CO3 solution, the degummed fibroin fiber could be dissolved in highly concentrated neutral salts such as CaCl2. The partially degraded polypeptides of silk fibroin, commonly called as regenerated liquid silk, could be obtained via water dialysis. The silk fibroin ...
    A Method for SUMO Modification of Proteins in vitro
    Authors:  Christine C. Lee, Bing Li, Hongtao Yu and Michael J. Matunis, date: 10/05/2018, view: 1494, Q&A: 0
    [Abstract] The Small Ubiquitin-related Modifier (SUMO) is a protein that is post-translationally added to and reversibly removed from other proteins in eukaryotic cells. SUMO and enzymes of the SUMO pathway are well conserved from yeast to humans and SUMO modification regulates a variety of essential cellular processes including transcription, chromatin ...
    Identifying Protein Interactions with Histone Peptides Using Bio-layer Interferometry
    Authors:  Bingbing Ren, Ahmed Mahmoud Mohammed Sayed, Hwei Ling Tan, Yu Keung Mok and Ee Sin Chen, date: 09/20/2018, view: 1719, Q&A: 0
    [Abstract] Histone post-translational modifications (PTMs) regulate numerous cellular processes, including gene transcription, cell division, and DNA damage repair. Most histone PTMs affect the recruitment or exclusion of reader proteins from chromatin. Here, we present a protocol to measure affinity and interaction kinetics between histone peptides and the ...
    Characterization of Protein Domain Function via in vitro DNA Shuffling
    Authors:  Kathy Hiu Laam Po, Edward Wai Chi Chan and Sheng Chen, date: 06/05/2018, view: 1584, Q&A: 0
    [Abstract] We recently investigated the molecular events that drive evolution of the CTX-M-type β-lactamases by DNA shuffling of fragments of the blaCTX-M-14 and blaCTX-M-15 genes. Analysis of a total of 51 hybrid enzymes showed that enzymatic activity could be maintained in most cases, yet the enzymatically active ...
    Histone Deubiquitination Assay in Nicotiana benthamiana
    Authors:  Shujing Liu and Lars Hennig, date: 03/05/2018, view: 1556, Q&A: 0
    [Abstract] Histone modifications are a group of post-translational modifications on histones which can alter chromatin structure and affect gene expression. Histone ubiquitination is a histone modification found in particular on histone H2A and H2B. Histone ubiquitination can be reversed by ubiquitin-specific proteases (UBP). Here, we describe an in vivo ...
    In vitro AMPylation Assays Using Purified, Recombinant Proteins
    Authors:  Matthias C. Truttmann and Hidde L. Ploegh, date: 07/20/2017, view: 3095, Q&A: 0
    [Abstract] Post-translational protein modifications (PTMs) orchestrate the activity of individual proteins and ensure their proper function. While modifications such as phosphorylation or glycosylation are well understood, more unusual modifications, including nitrosylation or AMPylation remain comparatively poorly characterized. Research on protein ...
    Phos-tag Immunoblot Analysis for Detecting IRF5 Phosphorylation
    Authors:  Go R. Sato, Tatsuma Ban and Tomohiko Tamura, date: 05/20/2017, view: 6324, Q&A: 0
    [Abstract] While the activation of the transcription factor interferon regulatory factor 5 (IRF5) is critical for the induction of innate immune responses, it also contributes to the pathogenesis of the autoimmune disease systemic lupus erythematosus (SLE). IRF5 phosphorylation is a hallmark of its activation in the Toll-like receptor (TLR) pathway, where ...
    Semi-quantitative Analysis of H4K20me1 Levels in Living Cells Using Mintbody
    Authors:  Yuko Sato and Hiroshi Kimura, date: 05/20/2017, view: 4348, Q&A: 0
    [Abstract] Eukaryotic nuclear DNA wraps around histone proteins to form a nucleosome, a basic unit of chromatin. Posttranslational modification of histones plays an important role in gene regulation and chromosome duplication. Some modifications are quite stable to be an epigenetic memory, and others exhibit rapid turnover or fluctuate during the cell cycle. ...



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