ArticlePloS one2010
Preparative scale production of functional mouse aquaporin 4 using different cell-free expression modes.
Article in PloS one, 2010. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 21 papers.
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Who cites it
21 citing papers in PubMed, 44 citations in OpenAlex.
- Applications of Cell-Free Synthesized Membrane Protein Precipitates.Methods in molecular biology (Clifton, N.J.) · 2022Article
- Review
- Functional Analysis of Aquaporin Water Permeability Using anFrontiers in bioengineering and biotechnology · 2020Article
- Article
- Quantification of Water Flux in Vesicular Systems.Scientific reports · 2018Article
- Cell-free protein synthesis enabled rapid prototyping for metabolic engineering and synthetic biology.Synthetic and systems biotechnology · 2018Review
- Coupling neutron reflectivity with cell-free protein synthesis to probe membrane protein structure in supported bilayers.Scientific reports · 2017Article
- The Water Permeability and Pore Entrance Structure of Aquaporin-4 Depend on Lipid Bilayer Thickness.Biophysical journal · 2016Article
- Functional characterization of p7 viroporin from hepatitis C virus produced in a cell-free expression system.Protein expression and purification · 2016Article
- Expression, Purification, and Characterization of a Sucrose Nonfermenting 1-Related Protein Kinases 2 ofBioMed research international · 2016Article
- Can Stabilization and Inhibition of Aquaporins Contribute to Future Development of Biomimetic Membranes?Membranes · 2015Review
- A refined model of water and CO₂ membrane diffusion: effects and contribution of sterols and proteins.Scientific reports · 2014Article
- High-level soluble expression of one model olfactory receptor (ODR-10) in Escherichia coli cell-free system.World journal of microbiology & biotechnology · 2014Article
- Aquaporin-4 expression in post-traumatic syringomyelia.Journal of neurotrauma · 2013Article
- The water permeability of lens aquaporin-0 depends on its lipid bilayer environment.Experimental eye research · 2013Article
- Water permeability of aquaporin-4 channel depends on bilayer composition, thickness, and elasticity.Biophysical journal · 2012Article
- Functional characterization of a novel aquaporin from Dictyostelium discoideum amoebae implies a unique gating mechanism.The Journal of biological chemistry · 2012Article
- Fluorescent in situ folding control for rapid optimization of cell-free membrane protein synthesis.PloS one · 2012Article
- Preparative scale cell-free production and quality optimization of MraY homologues in different expression modes.The Journal of biological chemistry · 2011Article
- Functional expression of the PorAH channel from Corynebacterium glutamicum in cell-free expression systems: implications for the role of the naturally occurring mycolic acid modification.The Journal of biological chemistry · 2011Article
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Authors and funding
8 authors at 3 institutions in 2 countries.
Funding
No grant is acknowledged in the PubMed record.
Abstract
The continuous progress in the structural and functional characterization of aquaporins increasingly attracts attention to study their roles in certain mammalian diseases. Although several structures of aquaporins have already been solved by crystallization, the challenge of producing sufficient amounts of functional proteins still remains. CF (cell free) expression has emerged in recent times as a promising alternative option in order to synthesize large quantities of membrane proteins, and the focus of this report was to evaluate the potential of this technique for the production of eukaryotic aquaporins. We have selected the mouse aquaporin 4 as a representative of mammalian aquaporins. The protein was synthesized in an E. coli extract based cell-free system with two different expression modes, and the efficiencies of two modes were compared. In both, the P-CF (cell-free membrane protein expression as precipitate) mode generating initial aquaporin precipitates as well as in the D-CF (cell-free membrane protein expression in presence of detergent) mode, generating directly detergent solubilized samples, we were able to obtain mg amounts of protein per ml of cell-free reaction. Purified aquaporin samples solubilized in different detergents were reconstituted into liposomes, and analyzed for the water channel activity. The calculated P(f) value of proteoliposome samples isolated from the D-CF mode was 133 µm/s at 10°C, which was 5 times higher as that of the control. A reversible inhibitory effect of mercury chloride was observed, which is consistent with previous observations of in vitro reconstituted aquaporin 4. In this study, a fast and convenient protocol was established for functional expression of aquaporins, which could serve as basis for further applications such as water filtration.
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