Protocols in Current Issue
    Expression and Purification of the Cas10-Csm Complex from Staphylococci
    Authors:  Lucy Chou-Zheng and Asma Hatoum-Aslan, date: 06/05/2017, view: 944, 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: 945, 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: 1203, 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: 1244, 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: 1300, 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: 1835, 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: 2007, 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 ...
    Preparation of Respiratory Syncytial Virus with High or Low Content of Defective Viral Particles and Their Purification from Viral Stocks
    Authors:  Yan Sun and Carolina B. López, date: 05/20/2016, view: 2893, Q&A: 0
    [Abstract] Respiratory syncytial virus (RSV) belongs to the paramyxovirus family that includes many clinically relevant viruses, such as the human metapneumovirus and measles. RSV infection can cause severe disease in infants, the elderly, and some immunocompromised adults. During RSV replication, a series of truncated forms of the viral genome are ...
    Purification of a Protein Exhibiting Isoleucine 2-epimerase Activity from Lactobacillus otakiensis JCM 15040
    Authors:  Yuta Mutaguchi and Toshihisa Ohshima, date: 10/20/2015, view: 2156, Q&A: 0
    [Abstract] Prominent accumulation of D-leucine, D-allo-isoleucine and D-valine was observed in the culture medium of the heterofermentative bacterial species, Lactobacillus otakiensis (L. otakiensis) JCM 15040. The racemase enzyme that resulted in this accumulation, isoleucine 2-epimerase, was purified from the bacterial cells. ...
    Purification of 70S Ribosomes from Bacillus subtilis
    Authors:  Shota Suzuki, Genki Akanuma and Fujio Kawamura, date: 04/05/2015, view: 3318, Q&A: 0
    [Abstract] The eubacterial ribosome (70S) is a macromolecular complex that is composed of a small (30S) subunit and a large (50S) subunit. The small subunit comprises the 16S ribosomal RNA (rRNA) and more than 20 ribosomal proteins (r-proteins), whereas the large subunit comprises the 23S and 5S rRNAs and more than 30 r-proteins. Bacillus subtilis ( ...