Chemistry and Physics Faculty Articles
Title
Ligand Induced Conformational Changes of a Membrane Transporter in E. coli Cells Observed with DEER/PELDOR
Document Type
Article
Publication Date
2-17-2016
Publication Title
Journal of the American Chemical Society
ISSN
0002-7863
Volume
138
Issue/No.
6
First Page
1844
Last Page
1847
Abstract
An unrealized goal in structural biology is the determination of structure and conformational change at high resolution for membrane proteins within the cellular environment. Pulsed electron–electron double resonance (PELDOR) is a well-established technique to follow conformational changes in purified membrane protein complexes. Here we demonstrate the first proof of concept for the use of PELDOR to observe conformational changes in a membrane protein in intact cells. We exploit the fact that outer membrane proteins usually lack reactive cysteines and that paramagnetic spin labels entering the periplasm are selectively reduced to achieve specific labeling of the cobalamin transporter BtuB in Escherichia coli. We characterize conformational changes in the second extracellular loop of BtuB upon ligand binding and compare the PELDOR data with high-resolution crystal structures. Our approach avoids detergent extraction, purification, and reconstitution usually required for these systems. With this approach, structure, function, conformational changes, and molecular interactions of outer membrane proteins can be studied at high resolution in the cellular environment.
Additional Comments
Deutsche Forschungsgemeinschaft grant #: SFB 807;; NIGMS grant #: GM035215
NSUWorks Citation
Joseph, B., Sikora, A., & Cafiso, D. S. (2016). Ligand Induced Conformational Changes of a Membrane Transporter in E. coli Cells Observed with DEER/PELDOR. Journal of the American Chemical Society, 138, (6), 1844 - 1847. https://doi.org/10.1021/jacs.5b13382. Retrieved from https://nsuworks.nova.edu/cnso_chemphys_facarticles/211
DOI
10.1021/jacs.5b13382
Comments
©2016 American Chemical Society