Protocols in Current Issue
    CENP-C Phosphorylation by CDK1 in vitro
    Authors:  Reito Watanabe, Masatoshi Hara, Mariko Ariyoshi and Tatsuo Fukagawa, date: 01/05/2021, view: 399, Q&A: 0

    Accurate chromosome segregation during mitosis requires the kinetochore, a large protein complex, which makes a linkage between chromosomes and spindle microtubes. An essential kinetochore component, CENP-C, is phosphorylated by Cyclin-B-Cyclin dependent kinase 1 (CDK1) that is a master kinase for mitotic progression, promoting proper kinetochore

    Isoelectric Focusing to Quantify Rhodopsin Phosphorylation in Mouse Retina
    Authors:  Sowmya Bekshe Lokappa, M. Carter Cornwall and Jeannie Chen, date: 07/20/2019, view: 3397, Q&A: 0
    [Abstract] Rhodopsin is a G-protein coupled receptor (GPCR) that mediates vision under dim light. Upon light exposure, rhodopsin is phosphorylated at multiple serine and threonine sites at its carboxyl-terminus by rhodopsin kinase (GRK1). This, in turn, reduces its ability to activate the visual G-protein transducin. Binding of light-activated, ...
    Isolation of Thylakoid Membranes from the Cyanobacterium Synechocystis sp. PCC 6803 and Analysis of Their Photosynthetic Pigment-protein Complexes by Clear Native-PAGE
    Authors:  Josef Komenda, Vendula Krynická and Tomas Zakar, date: 01/05/2019, view: 3950, Q&A: 0
    [Abstract] Cyanobacteria represent a frequently used model organism for the study of oxygenic photosynthesis. They belong to prokaryotic microorganisms but their photosynthetic apparatus is quite similar to that found in algal and plant chloroplasts. The key players in light reactions of photosynthesis are Photosystem I and Photosystem II complexes (PSI and ...
    Preparation of Cerebellum Granule Neurons from Mouse or Rat Pups and Evaluation of Clostridial Neurotoxin Activity and Their Inhibitors by Western Blot and Immunohistochemistry
    Authors:  Domenico Azarnia Tehran and Marco Pirazzini, date: 07/05/2018, view: 6651, Q&A: 0
    [Abstract] Cerebellar Granule Neurons (CGN) from post-natal rodents have been widely used as a model to study neuronal development, physiology and pathology. CGN cultured in vitro maintain the same features displayed in vivo by mature cerebellar granule cells, including the development of a dense neuritic network, neuronal activity, ...
    Detection of Catalase Activity by Polyacrylamide Gel Electrophoresis (PAGE) in Cell Extracts from Pseudomonas aeruginosa
    Authors:  Magdalena Pezzoni, Ramón A. Pizarro and Cristina S. Costa, date: 06/05/2018, view: 5290, Q&A: 0
    [Abstract] Bacteria in nature and as pathogens commonly face oxidative stress which causes damage to proteins, lipids and DNA. This damage is produced by the action of reactive oxygen species (ROS) such as hydrogen peroxide (H2O2), singlet oxygen, superoxide anion and hydroxyl radical. ROS are generated by antimicrobials, environmental ...
    Detection of Protein S-nitrosothiols (SNOs) in Plant Samples on Diaminofluorescein (DAF) Gels
    Authors:  Marta Rodríguez-Ruiz, Paulo T. Mioto, José M. Palma and Francisco J. Corpas , date: 09/20/2017, view: 4862, Q&A: 0
    [Abstract] In plant cells, the analysis of protein S-nitrosothiols (SNOs) under physiological and adverse stress conditions is essential to understand the mechanisms of Nitric oxide (NO)-based signaling. We adapted a previously reported protocol for detecting protein SNOs in animal systems (King et al., 2005) for plant samples. Briefly, ...
    Analyzing the Properties of Murine Intestinal Mucins by Electrophoresis and Histology
    Authors:  Ran Wang and Sumaira Z. Hasnain, date: 07/20/2017, view: 13882, Q&A: 0
    [Abstract] Specialized secretory cells known as goblet cells in the intestine and respiratory epithelium are responsible for the secretion of mucins. Mucins are large heavily glycosylated proteins and typically have a molecular mass higher than 106 Da. These large proteins are densely substituted with short glycan chains, which have many important ...
    Separation of Plant 6-Phosphogluconate Dehydrogenase (6PGDH) Isoforms by Non-denaturing Gel Electrophoresis
    [Abstract] 6-Phosphogluconate dehydrogenase (6PGDH; EC catalyzes the third and irreversible reaction of the pentose phosphate pathway (PPP). It carries out the oxidative decarboxylation of the 6-phosphogluconate to yield ribulose-5-phosphate, carbon dioxide and NADPH. In higher plants, 6PGDH has several subcellular localizations including cytosol, ...
    In Gel Kinase Assay
    Authors:  Gaston A. Pizzio and Pedro L. Rodriguez, date: 03/05/2017, view: 11170, Q&A: 0
    [Abstract] Proper spatiotemporal regulation of protein phosphorylation in cells and tissues is required for normal development and homeostasis. We present the protocol ‘In Gel Kinase Assay’, which is useful for protein kinase activity measurements from crude protein extracts. We have successfully used ‘In Gel Kinase Assay’ protocol to show that the ...
    Semi-denaturing Detergent Agarose Gel Electrophoresis (SDD-AGE)
    Authors:  Laura Molina-García and Fátima Gasset-Rosa, date: 11/20/2014, view: 15975, Q&A: 2
    [Abstract] Pathological proteins in neurodegenerative diseases suffer a conformational change to a misfolded amyloid state. Such pathological event leads to the aggregation of these proteins that indefinitely propagates as an altered form of itself, and harbor prion-like properties (Wickner, 1994; Prusiner, 2012). In addition to diseases, prions can also ...

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