Creative Biostructure provides unmatched custom Mempro™ P-Loop containing nucleoside triphosphate hydrolase (NTP) production services using detergent-free expression system.
P-loop containing NTP hydrolase superfamily, which contains common sequence patterns (two distinct motifs), the Walker A motif (the P-loop proper) and Walker B motif that binds, respectively, the beta and gamma phosphate moieties of the bound NTP, and a Mg2+ cation. Proteins from this superfamily share a P-loop NTP hydrolase fold, which is the most common domain of the several distinct nucleotide-binding protein folds. The biochemical reaction catalysed the P-loop NTP hydrolase fold is the hydrolysis of the beta-gamma phosphate bond of a bound NTP. Conventional approaches for P-loop containing NTP hydrolase purification need extraction from the membrane into detergent micelles. However, detergent can induce conformational change on membrane proteins. Our alternative detergent-free expression system can best reserve the native conformation of P-loop containing NTP hydrolases for further structural studies.
Figure 1. The structural model of Shikimate kinase. (OPM database)
Creative Biostructure has great experience in high-yield P-loop containing NTP hydrolase production using detergent-free membrane protein expression system, we can employ various strategies for Mempro™ detergent-free P-loop containing NTP hydrolase production, including:
Nanodiscs are the self-assembly system to stabilize membrane proteins removed from the membrane by membrane scaffold proteins (MSPs) in no need of detergent. Creative Biostructure can reconstitute P-loop containing NTP hydrolases into nanodiscs in detergent-free process: Expressing the P-loop containing NTP hydrolases in a cell-free system with the addition of pre-assembled nanodiscs.
The amphipathic amphipols have the ability to “trap” around transmembrane regions of proteins, allowing them to stay folded. Creative Biostructure can utilize A8-35 to solubilize P-loop containing NTP hydrolases during purification step.
The SMALPs are self-assembled by the simple addition of the SMA co-polymers. At neutral or alkaline pH, a disc-like structure assembles itself, encapsulating P-loop containing NTP hydrolases in a form amenable to be purified.
These novel detergent-free technologies for P-loop containing NTP hydrolase production can be obtained easily, and enabling more comprehensively structural and functional studies.
Creative Biostructure provides other various Mempro™ membrane protein production services. Please feel free to contact us for a detailed quote.
References:C. Tribet, et al. (1996). Amphipols: Polymers that keep membrane proteins soluble in aqueous solutions. Proc Natl Acad Sci U. S. A. 93: 15047-15050. J. Sang, et al. (2006). Duplication and combination of P-loop containing nucleotide triphosphate hydrolases superfamily. Wuhan University J. Nat. Sci., 11(3): 577-580. M. Jamshad, et al. (2011). Surfactant-free purification of membrane proteins with intact native membrane environment. Biochem. Soc. Trans., 39: 813-818.Y. Kawamura, et al. (2003). Systematic analyses of P-loop containing nucleotide triphosphate hydrolase superfamily based on sequence, structure and function. Genome Informatics, 14: 581-582.