Production of Membrane Proteins Through the Wheat-Germ Cell-Free Technology

2009 ◽  
pp. 213-218 ◽  
Author(s):  
Akira Nozawa ◽  
Hideaki Nanamiya ◽  
Yuzuru Tozawa
Keyword(s):  
Membrane Proteins ◽  
Germ Cell ◽  
Wheat Germ

Wheat Germ Cell-Free Overexpression for the Production of Membrane Proteins

2017 ◽  
pp. 91-108 ◽  
Author(s):  
Marie-Laure Fogeron ◽  
Aurélie Badillo ◽  
François Penin ◽  
Anja Böckmann
Keyword(s):  
Membrane Proteins ◽  
Germ Cell ◽  
Wheat Germ

Wheat germ cell-free expression: Two detergents with a low critical micelle concentration allow for production of soluble HCV membrane proteins

2015 ◽  
Vol 105 ◽  
pp. 39-46 ◽  
Author(s):  
Marie-Laure Fogeron ◽  
Aurélie Badillo ◽  
Vlastimil Jirasko ◽  
Jérôme Gouttenoire ◽  
David Paul ◽  
...  
Keyword(s):  
Membrane Proteins ◽  
Critical Micelle Concentration ◽  
Germ Cell ◽  
Wheat Germ ◽  
Free Expression

Wheat Germ Cell-Free Translation System as a Tool for In vitro Selection of Functional Proteins

2002 ◽  
Vol 5 (6) ◽  
pp. 473-480
Author(s):  
Bentham Science Publisher A.N. Alexandrov ◽  
Bentham Science Publisher V.Yu. Alakhov ◽  
Bentham Science Publisher A.I. Miroshnikov
Keyword(s):  
Germ Cell ◽  
Wheat Germ ◽  
In Vitro Selection ◽  
Translation System ◽  
Free Translation ◽  
Selection Of

scholarly journals The Quiescin Sulfhydryl Oxidase (hQSOX1b) Tunes the Expression of Resistin-Like Molecule Alpha (RELM-α or mFIZZ1) in a Wheat Germ Cell-Free Extract

PLoS ONE ◽  
2013 ◽  
Vol 8 (1) ◽  
pp. e55621 ◽  
Author(s):  
Wael Gad ◽  
Meera G. Nair ◽  
Karolien Van Belle ◽  
Khadija Wahni ◽  
Henri De Greve ◽  
...  
Keyword(s):  
Germ Cell ◽  
Wheat Germ ◽  
Cell Free Extract ◽  
Sulfhydryl Oxidase ◽  
Quiescin Sulfhydryl Oxidase

scholarly journals Mechanism of neutrophil chemiluminescence induced by wheat germ agglutinin: partial characterization of the antigens recognized by wheat germ agglutinin

Blood ◽  
1984 ◽  
Vol 64 (5) ◽  
pp. 1094-1102 ◽  
Author(s):  
Y Ozaki ◽  
J Iwata ◽  
T Ohashi
Keyword(s):  
Membrane Proteins ◽  
Wheat Germ Agglutinin ◽  
Wheat Germ ◽  
Sodium Dodecyl ◽  
Submaxillary Gland ◽  
Binding Property ◽  
Molecular Weights ◽  
Acetylneuraminic Acid ◽  
Cell Extracts ◽  
High Concentrations

Abstract Wheat germ agglutinin (WGA) stimulated neutrophils to produce significant levels of luminol-dependent chemiluminescence (CL). Since WGA is known to bind N-acetylglucosamine (GlcNAc) oligomers and N- acetylneuraminic acid (NANA), we attempted to determine which binding property of WGA is essential for induction of CL. The succinylated form of WGA (SuWGA), which is no longer able to bind NANA, was still able to induce CL. N-Acetylglucosamine at a concentration of 20 mmol/L almost completely inhibited WGA-induced CL production by neutrophils, whereas bovine submaxillary gland mucin, a potent blocker of NANA binding of WGA, failed to inhibit CL production. Lectins with the GlcNAc-binding property were examined for their ability to induce CL. Those that have higher valences and have a tendency to bind GlcNAc oligomers in the internal portion of glycoconjugates were able to induce CL, whereas those that have low valences and bind terminal GlcNAc of glycoconjugates failed to induce CL even at high concentrations. Attempts were made to characterize the neutrophil membrane proteins recognized by WGA. Glycoproteins with a molecular weight of 25,000 daltons were identified by a 50 mmol/L GlcNAc elution of WGA gels loaded with 125I-labeled neutrophil membrane proteins. Elution with 500 mumol/L GlcNAc trimer produced several glycoproteins of different molecular weights in addition to the glycoproteins of 25,000 daltons. 125I-labeled WGA and SuWGA were used for autoradiographic analysis of cell extracts of the neutrophils separated on sodium dodecyl sulfate polyacrylamide gels. WGA recognized multiple glycoproteins of different molecular weights, whereas SuWGA bound only a few of them. Glycoproteins of 25,000 daltons, probably corresponding to those identified by 50 mmol/L GlcNAc elution, were also recognized.

Malaria transmission-blocking vaccines: wheat germ cell-free technology can accelerate vaccine development

2019 ◽  
Vol 18 (10) ◽  
pp. 1017-1027
Author(s):  
Kazutoyo Miura ◽  
Mayumi Tachibana ◽  
Eizo Takashima ◽  
Masayuki Morita ◽  
Bernard N. Kanoi ◽  
...  
Keyword(s):  
Germ Cell ◽  
Malaria Transmission ◽  
Wheat Germ ◽  
Vaccine Development ◽  
Transmission Blocking ◽  
Transmission Blocking Vaccines

scholarly journals A novel family of expression vectors with multiple affinity tags for wheat germ cell-free protein expression

2020 ◽  
Vol 20 (1) ◽  
Author(s):  
Szilvia Krisztina Nagy ◽  
Brigitta Margit Kállai ◽  
Judit András ◽  
Tamás Mészáros
Keyword(s):  
Protein Expression ◽  
Germ Cell ◽  
Cleavage Site ◽  
Wheat Germ ◽  
In Vitro Translation ◽  
Affinity Tags ◽  
Free Protein ◽  
Target Proteins ◽  
Interaction Studies

Abstract Background Cell-free protein expression has become a widely used alternative of in vivo, cell-based systems in functional and structural studies of proteins. The wheat germ-based method outstands from the commercially available eukaryotic in vitro translation systems by its flexibility, high translation efficiency and success rate of properly folded eukaryotic protein synthesis. The original T7 promoter containing pEU3-NII vector was improved previously by addition of a ligation-independent cloning site, His6- and GST-tags, and a TEV protease cleavage site to facilitate the creation of recombinant plasmids, permit affinity purification, and enable production of purified, tag-free target proteins, respectively. Results Here, we describe a further development of pEU3-NII vector by inserting the rare-cutting, NotI restriction enzyme cleavage site to simplify vector linearization step prior to in vitro transcription. Additionally, His12, FLAG, and Halo affinity tag coding vectors have been created to increase detection sensitivity, specificity of interaction studies, and provide covalently linkable ligands for pull-down assays, respectively. Finally, the presented GST-His6, and GST-biotin double-tagging vectors could broaden the range of possibilities of protein-protein interaction studies. Conclusions The new generation of pEU3-NII vector family allows a more rapid production of translationally active mRNA and wheat germ cell-free expression of target proteins with a wide variety of affinity tags thus enables designing flexible and diverse experimental arrangement for in vitro studies of proteins.

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