Protocols in Current Issue
    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: 1126, 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 ...
    Selective Isolation of Retroviruses from Extracellular Vesicles by Intact Virion Immunoprecipitation
    Authors:  Tyler Milston Renner, Kasandra Bélanger and Marc-André Langlois, date: 09/05/2018, view: 1309, Q&A: 0
    [Abstract] There exists a wide variety of techniques to isolate and purify viral particles from cell culture supernatants. However, these techniques vary greatly in ease of use, purity, yield and impact on viral structural integrity. Most importantly, it is becoming evident that secreted extracellular vesicles (EVs) co-purify with retroviruses using nearly ...
    Easy and Efficient Permeabilization of Cyanobacteria for in vivo Enzyme Assays Using B-PER
    Authors:  Simon Matthé Erstad and Yumiko Sakuragi, date: 01/05/2018, view: 2274, Q&A: 0
    [Abstract] Cyanobacteria are photosynthetic bacteria that thrive in diverse ecosystems and play major roles in the global carbon cycle. The abilities of cyanobacteria to fix atmospheric CO2 and to allocate the fixed carbons to chemicals and biofuels have attracted growing attentions as sustainable microbial cell factories. A better understanding ...
    Expression and Purification of the Cas10-Csm Complex from Staphylococci
    Authors:  Lucy Chou-Zheng and Asma Hatoum-Aslan, date: 06/05/2017, view: 4916, Q&A: 0
    [Abstract] CRISPR-Cas (Clustered regularly interspaced short palindromic repeats-CRISPR-associated proteins) is a class of prokaryotic immune systems that degrade foreign nucleic acids in a sequence-specific manner. These systems rely upon ribonucleoprotein complexes composed of Cas nucleases and small CRISPR RNAs (crRNAs). Staphylococcus epidermidis ...
    Isolation of Ribosomal Particles from the Unicellular Cyanobacterium Synechocystis sp. PCC 6803
    Authors:  Carla V. Galmozzi and M. Isabel Muro-Pastor , date: 03/20/2017, view: 3727, Q&A: 0
    [Abstract] Isolation of ribosomal particles is an essential step in the study of ribosomal components as well as in the analysis of trans-acting factors that interact with the ribosome to regulate protein synthesis and modulate the expression profile of the cell in response to different environmental conditions. In this protocol, we describe a ...
    Expression and Purification of Cyanobacterial Circadian Clock Protein KaiC and Determination of Its Auto-phosphatase Activity
    Authors:  Qiang Chen, Lingling Yu, Xiao Tan and Sen Liu, date: 02/20/2017, view: 4038, Q&A: 0
    [Abstract] Circadian rhythms are biological processes displaying an endogenous oscillation with a period of ~24 h. They allow organisms to anticipate and get prepared for the environmental changes caused mainly by the rotation of Earth. Circadian rhythms are driven by circadian clocks that consist of proteins, DNA, and/or RNA. Circadian clocks of ...
    Production, Purification and Crystallization of a Prokaryotic SLC26 Homolog for Structural Studies
    Authors:  Yung-Ning Chang, Farooque R. Shaik, Yvonne Neldner and Eric R. Geertsma, date: 02/05/2017, view: 4232, Q&A: 0
    [Abstract] The SLC26 or SulP proteins constitute a large family of anion transporters that are ubiquitously expressed in pro- and eukaryotes. In human, SLC26 proteins perform important roles in ion homeostasis and malfunctioning of selected members is associated with diseases. This protocol details the production and crystallization of a prokaryotic SLC26 ...
    Protein Expression Protocol for an Adenylate Cyclase Anchored by a Vibrio Quorum Sensing Receptor
    Authors:  Stephanie Beltz and Joachim E. Schultz, date: 01/20/2017, view: 4421, Q&A: 0
    [Abstract] The direct regulation of a mycobacterial adenylate cyclase (Rv1625c) via exchange of its membrane anchor by the quorum sensing receptor CqsS (Vibrio harveyi) has recently been reported (Beltz et al., 2016). This protocol describes the expression and membrane preparation for these chimeric proteins.
    Heterologous Expression and Purification of the Magnesium Transporter A (MgtA) in Escherichia coli
    Authors:  Saranya Subramani and Jens Preben Morth, date: 11/20/2016, view: 5041, Q&A: 0
    [Abstract] The magnesium transporter A (MgtA) is a magnesium transporting P-type ATPase present in prokaryotes and plants (Subramani et al., 2016). In Salmonella typhimurium and Escherichia coli (E. coli), MgtA is expressed only in magnesium limiting conditions and plays an important role in Mg2+ homeostasis ...
    Purification and Identification of Novel Host-derived Factors for Influenza Virus Replication from Human Nuclear Extracts
    Authors:  Kenji Sugiyama and Kyosuke Nagata, date: 09/20/2016, view: 5213, Q&A: 0
    [Abstract] Recently, we identified two host cell-derived proteins as novel stimulatory factors of influenza virus RNA replication process, termed “Influenza virus REplication Factor-2 (IREF-2)”, from human nuclear extracts (NEs) by employing biochemical complementation assays (Sugiyama et al., 2015). Herein, we describe detailed methods for ...

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