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Mempro™ Cell-Free PX Domain-Containing Protein Production

Creative Biostructure can supply customized Mempro™ cell-free phox homology (PX) domain-containing protein production services. You can count on us all through your project.

Mempro™ cell-free protein production system is the innovative systems to produce integral membrane proteins such as G-protein coupled receptors (GPCRs), which can bypass the limitations that encountered in cell-based protein production system, such as host cell toxicity, low yield and solubilization and purification using detergents. PX domain is an evolutionarily conserved phospholipid-binding protein domain of about 120 amino acids. The p40phox and p47phox derived from the NADPH oxidase complex belong to PX domain-containing proteins. The PX domain has been shown to be involved in membrane trafficking, including nexins, phospholipase D, and phosphoinositide-3-kinases, which can bind primarily phosphatidylinositol 3-phosphate [PtdIns(3)P] lipids. The PX domain-containing proteins can interact with other domains and proteins.

MemproTM Cell-Free PX Domain-Containing Protein Production Figure 1. Biological function of PX domain proteins. (Nature Cell Biol., 2001)

Creative Biostructure has strong expertise in high-yield PX domain-containing protein production using cell-free protein expression systems, we can provide various strategies for native activity of desired membrane proteins, including:

E. coli S30 extract is popularly used for PX domain protein production due to inexpensive culture media. Wheat germ extract can provide eukaryotic translation environment for PX domain protein expression. Baculovirus expression system from Autographa californica nucleopolyhedrovirus (AcNPV) is also a major tool for PX domain protein expression, which is developed in the early stage that transfected insect cells in combination with vectors derived from the baculovirus. Crude rabbit reticulocyte lysate is also the most widely used for eukaryotic cell-free membrane protein expression system. To select an appropriate source for protein expression, many factors, such as the availability of materials, protein origin and complexity, downstream processing, etc., should be considered.

With the Mempro™ cell-free protein production platform, Creative Biostructure is capable of expressing, purifying and crystallizing PX domain proteins to facilitate the study of their biological functions. Mempro™ cell-free membrane protein production has a wide range of advantages, including:

  • Very high yield;
  • Low cost in time and money;
  • Lack of toxicity;
  • Incorporation of non-natural amino acids into protein structure;
  • Relatively tolerant of additives.

Notably, One challenge associated with Mempro™ cell-free protein production is that degradation of the DNA by endogenous nucleases. Creative Biostructure can optimize Mempro™ cell-free protein expression system using plasmids owing to they are circular and have no end for the exonucleases to attach.

We provide other various Mempro™ membrane protein production services. Please feel free to contact us for a detailed quote.

References:
A. Simonsen and H. Stenmark (2001). PX domains: attracted by phosphoinositides. Nature Cell Biol., 3: E179 - E182.
Cell-free protein synthesis. (https://en.wikipedia.org/wiki/Cell-free_protein_synthesis)
F. Bernhard, et al. (2013). Cell-free expression - making a mark. Cur. Opin. Struct. Biol., 23: 374-380.
L. F.Seet and W. Hong (2006). The Phox (PX) domain proteins and membrane traffic. Biochim. Biophy. Acta , 1761(8): 878-896.

PX domain. (https://en.wikipedia.org/wiki/PX_domain)

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