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Mempro™ Cell-Free Cytochrome P450 Production

Creative Biostructure can provide customized Mempro™ protein production services for cytochrome P450 (CYP) superfamily proteins using cell-free expression system. Our cell-free expression systems can preserve the native activity of CYPs for your specific projects.

Mempro™ cell-free protein production system is the innovative system to produce intergral membrane proteins (including GPCRs and CYPs) which can bypass the limitations that encountered in cell-based protein production systems, such as host cell toxicity, low yield and solubilization and purification using detergents. CYPs, also called hemoproteins, are composed of a superfamily of similar proteins containing a hemo cofactor. CYPs is the key liver endoplasmic reticulum protein that playing an important role in drug metabolism (Figure 1).

Mempro™ Cell-Free Cytochrome P450 Production Figure 1. Potential routes to cytochrome P450 biocatalysts. (Trends Biotechnol., 2011)

Creative Biostructure has a wealth of experience in high-yield cytochrome P450 production using cell-free protein expression systems, we can provide various strategies for Mempro™ cell-free protein production, including:

Among Mempro™ cell-free protein production systems, E. coli S30 extract is commonly used for CYPs production. Wheat germ extract can provide eukaryotic translation system for CYPs expression. Baculovirus expression system from Autographa californica nucleopolyhedrovirus (AcNPV) is also a major system for CYPs 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 integral membrane protein expression systems. To adopt an appropriate source of the extract, 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 CYPs to facilitate the study of their biological functions. Mempro™ cell-free protein production has a large 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.

Most notably, One challenge associated with Mempro™ cell-free membrane protein production is that degradation of the DNA by endogenous nucleases. Creative Biostructure can optimize Mempro™ cell-free membrane protein production for cytochrome P450 using plasmids due to they are circular and have no end for the exonucleases to attach.

Creative Biostructure provides other various Mempro™ membrane protein production services. Please feel free to contact us for a detailed quote.

References:
V. B. , Urlacher and M. Girhard (2011). Cytochrome P450 monooxygenases: an update on perspectives for synthetic application. Trends Biotechnol., 30(1): 26-36.
F. Bernhard, et al. (2013). Cell-free expression - making a mark. Cur. Opin. Struct. Biol., 23: 374-380.
K. S. Bhat and G. Padmanaban (1978). Cytochrome P-450 synthesis in vivo and in a cell-free system from rat liver. FEBS Lett., 89(2): 337-340.


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