[NMR paper] NMR-based Conformational Ensembles Explain pH-Gated Opening and Closing of OmpG Channel.

[nmrlearner]

NMR-based Conformational Ensembles Explain pH-Gated Opening and Closing of OmpG Channel.

Related Articles NMR-based Conformational Ensembles Explain pH-Gated Opening and Closing of OmpG Channel.

J Am Chem Soc. 2013 Sep 10;

Authors: Zhuang T, Chisholm C, Chen M, Tamm LK

Abstract
The outer membrane protein G (OmpG) is a monomeric 33 kDa 14-stranded ?-barrel membrane protein functioning as a non-specific porin for the uptake of oligosaccharides in E. coli. Two different crystal structures of OmpG obtained at different values of pH suggest a pH-gated pore opening mechanism. In these structures, extracellular loop 6 extends away from the barrel wall at neutral pH, but is folded back into the pore lumen at low pH, blocking transport through the pore. Loop 6 was invisible in a previously published solution NMR structure of OmpG in DPC micelles, presumably due to conformational exchange on an intermediate NMR time-scale. Here we present an NMR paramagnetic relaxation enhancement (PRE)-based approach to visualize the conformational dynamics of loop 6 and to calculate conformational ensembles that explain the pH-gated opening and closing of the OmpG channel. The different loop conformers detected by the PRE ensemble calculations were validated by disulfide cross-linking of strategically engineered cysteines and electrophysiological single channel recordings. The results indicate a more dynamically regulated channel opening and closing than previously thought and reveal additional membrane-associated conformational ensembles at pH 6.3 and 7.0. We anticipate this approach to be generally applicable to detect and characterize functionally important conformational ensembles of membrane proteins.


PMID: 24020969 [PubMed - as supplied by publisher]



More...