Issue 16, 2019, Issue in Progress
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RSC Advances

Doubly spin-labeled nanodiscs to improve structural determination of membrane proteins by ESR†

Chieh-Chin Li, ORCID logo a   Chien-Lun Hung, ORCID logo a   Pei-Shan Yeh,a   Chi-En Lia  and  Yun-Wei Chiang ORCID logo *a  

* Corresponding authors

a Department of Chemistry, Frontier Research Center on Fundamental and Applied Sciences of Matters, National Tsing Hua University, Hsinchu 30013, Taiwan
E-mail: ywchiang@gapp.nthu.edu.tw

Abstract

Pulsed dipolar spectroscopy (PDS) is a powerful tool to explore conformational changes of membrane proteins (MPs). However, the MPs suffer from relatively weak dipolar signals due to their complex nature in membrane environments, which consequently reduces the interspin distance resolution obtainable by PDS. Here we report the use of nanodiscs (NDs) to improve the distance resolution. Two genetically engineered membrane scaffold protein mutants are introduced, each of which is shown to form double-labeled ND efficiently and with high homogeneity. The resultant interspin distance distribution is featured by a small distribution width, suggesting high resolution. When PDS is performed on a binary mixture of the double-labeled ND devoid of MPs and the un-labeled ND with incorporated double-labeled MPs, the overall amplitude of dipolar signals is increased, leading to a critical enhancement of the distance resolution. A theoretical foundation is provided to validate the analysis. With this approach, the determination of MP structures can be studied at high resolution in NDs.

Graphical abstract: Doubly spin-labeled nanodiscs to improve structural determination of membrane proteins by ESR

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Article information

DOI
https://doi.org/10.1039/C9RA00896A
Article type
Paper
Submitted
02 Feb 2019
Accepted
14 Mar 2019
First published
19 Mar 2019
This article is Open Access
Creative Commons BY-NC license

RSC Adv., 2019,9, 9014-9021

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Doubly spin-labeled nanodiscs to improve structural determination of membrane proteins by ESR

C. Li, C. Hung, P. Yeh, C. Li and Y. Chiang, RSC Adv., 2019, 9, 9014 DOI: 10.1039/C9RA00896A

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