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    Detection of Membrane Protein Interactions by Cell-based Tango Assays
    [Abstract] The Tango assay is a protein-protein interaction assay, in which a transcription factor (rTA) is fused to a membrane-bound protein via a linker that contains a cleavage site for TEV protease, whereas a soluble interaction partner is fused to TEV protease (Barnea et al., 2008). Association between the two interaction partners leads to an ...
    Protein Expression and Purification of the Hsp90-Cdc37-Cdk4 Kinase Complex from Saccharomyces cerevisiae
    Authors:  Kliment A. Verba and David A. Agard, date: 10/05/2017, view: 1017, Q&A: 0
    [Abstract] Interactions between Hsp90, its co-chaperone Cdc37 and kinases have been biochemically studied for over three decades and have been shown to be functionally important in organisms from yeast to humans. However, formation of a stable complex for structural studies has been elusive. In this protocol we describe expression and purification of ...
    Expression and Purification of a Mammalian P2X7 Receptor from Sf9 Insect Cells
    Authors:  Akira Karasawa and Toshimitsu Kawate, date: 09/05/2017, view: 1427, Q&A: 0
    [Abstract] The P2X7 receptor is an extracellular ATP-gated ion channel found only in eukaryotes (Bartlett et al., 2014). Due to its unique properties among P2X receptors, such as formation of a large conductance pore, the P2X7 receptor has been implicated in devastating diseases like chronic pain (North and Jarvis, 2013). However, mechanisms ...
    Expression and Purification of Mini G Proteins from Escherichia coli
    Authors:  Byron Carpenter and Christopher G. Tate, date: 04/20/2017, view: 2322, Q&A: 0
    [Abstract] Heterotrimeric G proteins modulate intracellular signalling by transducing information from cell surface G protein-coupled receptors (GPCRs) to cytoplasmic effector proteins. Structural and functional characterisation of GPCR–G protein complexes is important to fully decipher the mechanism of signal transduction. However, native G proteins are ...
    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: 1914, 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 ...
    Expression and Purification of the GRAS Domain of Os-SCL7 from Rice for Structural Studies
    Authors:  Shengping Li, Yanhe Zhao and Yunkun Wu, date: 02/05/2017, view: 1798, Q&A: 0
    [Abstract] GRAS proteins, named after the first three members GAI, RGA and SRC, has been found in 294 embryophyta species and is represented by 1,035 sequences. They belong to a plant-specific protein family and play essential roles in plant growth and development. Proteins in this family are defined as minimally containing a conserved GRAS domain, which is ...
    Expression, Purification and Crystallization of Recombinant Arabidopsis Monogalactosyldiacylglycerol Synthase (MGD1)
    Authors:  Joana Rocha, Valerie Chazalet and Christelle Breton, date: 12/20/2016, view: 1818, Q&A: 0
    [Abstract] In plant cells, galactolipids are predominant, representing up to 50% of the lipid content in photosynthetic tissues. Galactolipid synthesis is initiated by MGDG synthases (MGDs), which use UDP-galactose as a donor sugar and diacylglycerol (DAG) as acceptor, to form monogalactosyldiacylglycerol (MGDG). This protocol is used to produce a ...
    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: 2676, 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 ...
    Expression, Purification and Crystallization of the Herpesvirus Nuclear Egress Complex (NEC)
    Authors:  Janna M. Bigalke and Ekaterina E. Heldwein, date: 07/20/2016, view: 2075, Q&A: 0
    [Abstract] The protocol describes the production and crystallization of the soluble form of the nuclear egress complex (NEC) from Herpes simplex virus 1 and Pseudorabies virus. The NEC is a heterodimer that consists of conserved proteins UL31 and UL34. NEC oligomerization deforms the inner nuclear membrane around the capsid in infected cells, thereby ...
    Preparation of Recombinant Galectin-3 for Cancer Studies
    Authors:  Kari Tyler, Sok-Hyong Lee and Erwin G. Van Meir, date: 01/05/2016, view: 3060, Q&A: 0
    [Abstract] Galectin-3 is a member of a class of proteins termed Galectins, characterized by their ability to bind glycans containing β-galactose (Cummings and Liu, 2009). Galectin-3 binds preferentially to proteoglycans terminating with N-acetyllactosamine (LacNAc) chains (i.e., tandem repeats of galactose) (Newlaczyl and Yu, 2011). Galectin-3 is ...