Biochemistry

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    Protocols in Current Issue
    Sulfatase Assay to Determine Influence of Plants on Microbial Activity in Soil
    Authors:  Anna Koprivova, Achim Schmalenberger and Stanislav Kopriva, date: 01/20/2020, view: 250, Q&A: 0
    Sulfatase activity is often used as a measure of the activity of soil microorganisms. It is thus a suitable tool to investigate the response of microbes to plants. Here we present a method to determine the influence of various Arabidopsis ...
    Protocols for Processing and Interpreting cryoEM Data Using Bsoft: A Case Study of the Retinal Adhesion Protein, Retinoschisin
    Author:  J. Bernard Heymann, date: 01/20/2020, view: 107, Q&A: 0
    The goal of cryoEM is to determine the structures of biomolecules from electron micrographs. In many cases the processing is straightforward and can be handled with routine protocols. In other cases, the properties and behavior of the specimen ...
    A Radioactive-free Kinase Inhibitor Discovery Assay Against the Trypanosoma brucei Glycogen Synthase Kinase-3 short (TbGSK-3s)
    Authors:  Antonia Efstathiou and Despina Smirlis, date: 01/20/2020, view: 83, Q&A: 0
    The identification of small molecules possessing inhibitory activity in vitro, against a given target kinase, is the first step in the drug discovery process. Herein, we describe a non radioactive protocol using luciferase-based ATP assay ...
    Studying Protein Aggregation in the Context of Liquid-liquid Phase Separation Using Fluorescence and Atomic Force Microscopy, Fluorescence and Turbidity Assays, and FRAP
    Authors:  W. Michael Babinchak and Witold K. Surewicz, date: 01/20/2020, view: 175, Q&A: 0
    Liquid-liquid phase separation (LLPS) underlies the physiological assembly of many membrane-less organelles throughout the cell. However, dysregulation of LLPS may mediate the formation of pathological aggregates associated with neurodegenerative ...
    Ribonucleoprotein Immunoprecipitation (RIP) Analysis
    Authors:  Jennifer L. Martindale, Myriam Gorospe and Maria L. Idda, date: 01/20/2020, view: 217, Q&A: 0
    RNAs and RNA-binding proteins (RBPs) can interact dynamically in ribonucleoprotein (RNP) complexes that play important roles in controlling gene expression programs. One of the powerful ways to investigate changes in the association of RNAs with an ...
    Sulfatase Assay to Determine Influence of Plants on Microbial Activity in Soil
    Authors:  Anna Koprivova, Achim Schmalenberger and Stanislav Kopriva, date: 01/20/2020, view: 250, Q&A: 0
    [Abstract] Sulfatase activity is often used as a measure of the activity of soil microorganisms. It is thus a suitable tool to investigate the response of microbes to plants. Here we present a method to determine the influence of various Arabidopsis genotypes on the function of soil microbiota using the sulfatase as a quantitative measure. We grew ...
    Protocols for Processing and Interpreting cryoEM Data Using Bsoft: A Case Study of the Retinal Adhesion Protein, Retinoschisin
    Author:  J. Bernard Heymann, date: 01/20/2020, view: 107, Q&A: 0
    [Abstract] The goal of cryoEM is to determine the structures of biomolecules from electron micrographs. In many cases the processing is straightforward and can be handled with routine protocols. In other cases, the properties and behavior of the specimen require adaptions to properly interpret the data. Here I describe the protocols for examining the higher ...
    A Radioactive-free Kinase Inhibitor Discovery Assay Against the Trypanosoma brucei Glycogen Synthase Kinase-3 short (TbGSK-3s)
    Authors:  Antonia Efstathiou and Despina Smirlis, date: 01/20/2020, view: 83, Q&A: 0
    [Abstract] The identification of small molecules possessing inhibitory activity in vitro, against a given target kinase, is the first step in the drug discovery process. Herein, we describe a non radioactive protocol using luciferase-based ATP assay for the identification of inhibitors for the short isoform of the Trypanosoma brucei’s ...
    Studying Protein Aggregation in the Context of Liquid-liquid Phase Separation Using Fluorescence and Atomic Force Microscopy, Fluorescence and Turbidity Assays, and FRAP
    Authors:  W. Michael Babinchak and Witold K. Surewicz, date: 01/20/2020, view: 175, Q&A: 0
    [Abstract] Liquid-liquid phase separation (LLPS) underlies the physiological assembly of many membrane-less organelles throughout the cell. However, dysregulation of LLPS may mediate the formation of pathological aggregates associated with neurodegenerative diseases. Here, we present complementary experimental approaches to study protein aggregation within ...
    Ribonucleoprotein Immunoprecipitation (RIP) Analysis
    Authors:  Jennifer L. Martindale, Myriam Gorospe and Maria L. Idda, date: 01/20/2020, view: 217, Q&A: 0
    [Abstract] RNAs and RNA-binding proteins (RBPs) can interact dynamically in ribonucleoprotein (RNP) complexes that play important roles in controlling gene expression programs. One of the powerful ways to investigate changes in the association of RNAs with an RBP of interest is by immunoprecipitation (IP) analysis of native RNPs. RIP (RNP ...
    Steady-state and Flux-based Trehalose Estimation as an Indicator of Carbon Flow from Gluconeogenesis or Glycolysis
    Authors:  Ritu Gupta and Sunil Laxman, date: 01/05/2020, view: 232, Q&A: 0
    [Abstract] Trehalose (and glycogen) is a major storage carbohydrate in many cells, including S. cerevisiae. Typically, trehalose (a disaccharide of glucose) is synthesized and stored through gluconeogenesis. However, trehalose can also be made directly from glucose, if glucose-6-phosphate is channeled away from glycolysis or pentose phosphate ...
    Laser Capture Micro-dissection (LCM) of Neonatal Mouse Forebrain for RNA Isolation
    Authors:  Achira Roy, Mei Deng, Kimberly A. Aldinger, Ian A. GLass and Kathleen J. Millen, date: 01/05/2020, view: 287, Q&A: 0
    [Abstract] Precise and reproducible isolation of desired cell types or layers from heterogeneous tissues is crucial to analyze specific gene profiles and molecular interactions in vivo. Forebrain is the core site of higher functions, like cognition and memory consolidation. It is composed of heterogeneous and distinct cell types, interconnected to ...
    Measurement of Chloroplastic NAD Kinase Activity and Whole Tissue NAD Kinase Assay
    Authors:  Yuuma Ishikawa, Maki Kawai-Yamada and Shin-nosuke Hashida, date: 01/05/2020, view: 249, Q&A: 0
    [Abstract] Nicotinamide adenine dinucleotide phosphate (NADP) synthesis requires nicotinamide adenine dinucleotide (NAD) kinase activity, substrate NAD and ATP. The NAD kinase responds to various environmental stimuli and its activity is regulated via various regulatory pathways, such as Ca2+-dependent and redox-dependent signals. Conventional in ...
    Analysis of Monosaccharides from Arabidopsis Seed Mucilage and Whole Seeds Using HPAEC-PAD
    Authors:  Gillian H. Dean, Kresimir Sola, Faride Unda, Shawn D. Mansfield and George W. Haughn, date: 12/20/2019, view: 400, Q&A: 0
    [Abstract] Arabidopsis seed coat epidermal cells deposit a significant quantity of mucilage, composed of the cell wall components pectin, hemicellulose, and cellulose, into the apoplast during development. When mature seeds are hydrated, mucilage extrudes to form a gelatinous capsule around the seed. Determining the monosaccharide composition of ...
    Semi-quantitative Determination of Protein Expression Using Immunohistochemistry Staining and Analysis: An Integrated Protocol
    Authors:  Alexandra R Crowe and Wei Yue, date: 12/20/2019, view: 722, Q&A: 0
    [Abstract] Semi-quantitative IHC is a powerful method for investigating protein expression and localization within tissues. The semi-quantitative immunohistochemistry (IHC) involves using software such as free software ImageJ Fiji to conduct deconvolution and downstream analysis. Currently, there is lack of an integrated protocol that includes a detailed ...



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