Optimization of the Production of Covalently Circularized Nanodiscs and Their Characterization in Physiological Conditions

Langmuir : the ACS Journal of Surfaces and Colloids
Yustina YusufTsyr-Yan Yu

Abstract

Lipid nanodiscs are widely used platforms for studying membrane proteins in a near-native environment. Lipid nanodiscs made with membrane scaffold proteins (MSPs) in the linear form have been well studied. Recently, a new kind of nanodisc made with MSPs in the circular form, referred to as covalently circularized nanodiscs (cNDs), has been reported to have some possible advantages in various applications. Given the potential of nanodisc technology, researchers in the field are very interested in learning more about this new kind of nanodisc, such as its reproducibility, production yield, and the possible pros and cons of using it. However, research on these issues is lacking. Here, we report a new study on nanodiscs made with circular MSPs, which are produced from a method different from the previously reported method. We show that our novel production method, detergent-assisted sortase-mediated ligation, can effectively avoid high-molecular-weight byproducts and also significantly improve the yield of the target proteins up to around 80% for larger circular MSP constructs. In terms of the application of circular MSPs, we demonstrate that they can be used to assemble nanodiscs using both synthetic lipids and native lipid extrac...Continue Reading

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Jun 28, 2018·Chembiochem : a European Journal of Chemical Biology·Jonas MiehlingFranz Hagn
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