scholarly journals Wheat germ in vitro translation to produce one of the most toxic sodium channel specific toxins

2014 ◽  
Vol 34 (4) ◽  
Author(s):  
Wael Gad ◽  
Rahma Ben-Abderrazek ◽  
Khadija Wahni ◽  
Didier Vertommen ◽  
Serge Muyldermans ◽  
...  
Keyword(s):  
Wheat Germ ◽  
Antibody Fragment ◽  
Scorpion Venom ◽  
Biologically Active ◽  
In Vitro Translation ◽  
Translation System ◽  
Single Domain Antibody ◽  
Venom Protein ◽  
Free Translation

A wheat germ embryo derived cell-free translation system expresses a biologically active, highly toxic scorpion venom protein that is fully neutralized by a camel single domain antibody fragment raised against the native scorpion toxin.

scholarly journals Gene expression in Acetabularia. I. Calibration of wheat germ cell-free translation system proteins as internal references for two-dimensional electrophoresis

1982 ◽  
Vol 58 (1) ◽  
pp. 23-33
Author(s):  
R.L. Shoeman ◽  
H.G. Schweiger
Keyword(s):  
Germ Cell ◽  
Isoelectric Point ◽  
Wheat Germ ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
In Vitro Translation ◽  
Translation System ◽  
Two Dimensional ◽  
Free Translation

Modification of existing two-dimensional techniques enables isoelectric focusing and sodium dodecyl sulphate polyacrylamide gel electrophoresis of complex mixtures of proteins to be completed within 8 h. The method was optimized to separate the protein components of a wheat germ cell-free translation system, providing a statistically proven resolution better than 0. 03 of a pH unit for the isoelectric point and 1000 for Mr. Fourteen of the more than 300 proteins separated were characterized with respect to Mr and isoelectric point relative to standard proteins under the same conditions. Stained wheat germ proteins thus serve as internal standards for analysis of in vitro translation products.

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 Towards a generic prototyping approach for therapeutically-relevant peptides and proteins in a cell-free translation system

2022 ◽  
Vol 13 (1) ◽  
Author(s):  
Yue Wu ◽  
Zhenling Cui ◽  
Yen-Hua Huang ◽  
Simon J. de Veer ◽  
Andrey V. Aralov ◽  
...  
Keyword(s):  
Interaction Analysis ◽  
Cost Effective ◽  
In Vitro Translation ◽  
Translation System ◽  
Activity Assessment ◽  
Free Translation ◽  
Macrocyclic Peptides ◽  
A Cell ◽  
Translation Systems

AbstractAdvances in peptide and protein therapeutics increased the need for rapid and cost-effective polypeptide prototyping. While in vitro translation systems are well suited for fast and multiplexed polypeptide prototyping, they suffer from misfolding, aggregation and disulfide-bond scrambling of the translated products. Here we propose that efficient folding of in vitro produced disulfide-rich peptides and proteins can be achieved if performed in an aggregation-free and thermodynamically controlled folding environment. To this end, we modify an E. coli-based in vitro translation system to allow co-translational capture of translated products by affinity matrix. This process reduces protein aggregation and enables productive oxidative folding and recycling of misfolded states under thermodynamic control. In this study we show that the developed approach is likely to be generally applicable for prototyping of a wide variety of disulfide-constrained peptides, macrocyclic peptides with non-native bonds and antibody fragments in amounts sufficient for interaction analysis and biological activity assessment.

Fast in vitro translation system immobilized on a surface via specific biotinylation of the ribosome

2008 ◽  
Vol 389 (9) ◽  
Author(s):  
Romualdas Stapulionis ◽  
Yuhong Wang ◽  
Graham T. Dempsey ◽  
Rama Khudaravalli ◽  
Karen Margrethe Nielsen ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Single Molecule ◽  
Dynamic Properties ◽  
Kinetic Studies ◽  
Biologically Active ◽  
In Vitro Translation ◽  
23S Rrna ◽  
Translation System ◽  
Peptide Bonds

Abstract The ribosome is the macromolecular machine responsible for translating the genetic code into polypeptide chains. Despite impressive structural and kinetic studies of the translation process, a number of challenges remain with respect to understanding the dynamic properties of the translation apparatus. Single-molecule techniques hold the potential of characterizing the structural and mechanical properties of macromolecules during their functional cycles in real time. These techniques often necessitate the specific coupling of biologically active molecules to a surface. Here, we describe a procedure for such coupling of functionally active ribosomes that permits single-molecule studies of protein synthesis. Oxidation with NaIO4 at the 3′ end of 23S rRNA and subsequent reaction with a biotin hydrazide produces biotinylated 70S ribosomes, which can be immobilized on a streptavidin-coated surface. The surface-attached ribosomes are fully active in poly(U) translation in vitro, synthesizing poly(Phe) at a rate of 3–6 peptide bonds/s per active ribosome at 37°C. Specificity of binding of biotinylated ribosomes to a streptavidin-coated quartz surface was confirmed by observation of individual fluorescently labeled, biotinylated 70S ribosomes, using total internal reflection fluorescence microscopy. Functional interactions of the immobilized ribosomes with various components of the protein synthesis apparatus are shown by use of surface plasmon resonance.

Preparation of a cell-free translation system from a wild-type strain of Neurospora crassa and translation of pyruvate kinase messenger RNA

1984 ◽  
Vol 4 (11) ◽  
pp. 979-986 ◽  
Author(s):  
M. Devchand ◽  
M. Kapoor
Keyword(s):  
Neurospora Crassa ◽  
Messenger Rna ◽  
In Vitro Translation ◽  
Translation System ◽  
Wild Type ◽  
Post Translational Modification ◽  
Globin Mrna ◽  
Free Translation ◽  
A Cell

A cell-free in vitro translation system exhibiting high activity has been developed from wild-type Neurospora crassa mycelium. The isolation is simple and fast, and the homogenization does not appear to affect the activity of mycelial proteases and nucleases. This system is capable of supporting efficient translation of exogenously added homologous RNA as demonstrated by the experiments with PK-specific mRNA. In addition, it translates heterologous RNA efficiently, shown by the translation of globin mRNA. We did not examine the Neurospora lysate for post-translational modification activity. The procedure used for the preparation of Neurospora cell-free extracts should be readily applicable to the other filamentous fungi.

Structural Determinants for Inhibitor Specificity and Selectivity in PDE2A Using the Wheat Germ in Vitro Translation System†

Biochemistry ◽  
2005 ◽  
Vol 44 (23) ◽  
pp. 8312-8325 ◽  
Author(s):  
André Iffland ◽  
Darcy Kohls ◽  
Simon Low ◽  
Jing Luan ◽  
Yan Zhang ◽  
...  
Keyword(s):  
Wheat Germ ◽  
In Vitro Translation ◽  
Translation System ◽  
Structural Determinants ◽  
Inhibitor Specificity

scholarly journals Development of a tRNA-dependent in vitro translation system

RNA ◽  
2001 ◽  
Vol 7 (5) ◽  
pp. 765-773 ◽  
Author(s):  
RICHARD J. JACKSON ◽  
SAWSAN NAPTHINE ◽  
IAN BRIERLEY
Keyword(s):  
In Vitro Translation ◽  
Translation System

scholarly journals Arabidopsis Cell-free Extract, ACE, a New In Vitro Translation System Derived from Arabidopsis Callus Cultures

2012 ◽  
Vol 53 (3) ◽  
pp. 602-602
Author(s):  
K. Murota ◽  
Y. Hagiwara-Komoda ◽  
K. Komoda ◽  
H. Onouchi ◽  
M. Ishikawa ◽  
...  
Keyword(s):  
Callus Cultures ◽  
In Vitro Translation ◽  
Translation System ◽  
Cell Free Extract ◽  
Arabidopsis Callus

scholarly journals An efficient in vitro translation system from mammalian cells lacking the translational inhibition caused by eIF2 phosphorylation

RNA ◽  
2008 ◽  
Vol 14 (3) ◽  
pp. 593-602 ◽  
Author(s):  
V. V. Zeenko ◽  
C. Wang ◽  
M. Majumder ◽  
A. A. Komar ◽  
M. D. Snider ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
In Vitro Translation ◽  
Translation System ◽  
Translational Inhibition ◽  
Eif2 Phosphorylation
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