Solid-state NMR structure determination of a membrane protein in <i>E. coli</i> cellular inner membrane.

Abstract

Structure determination of membrane proteins in native cellular membranes is critical to precisely reveal their structures in physiological conditions. However, it remains challenging for solid-state nuclear magnetic resonance (ssNMR) due to the low sensitivity and high complexity of ssNMR spectra of cellular membranes. Here, we present the structure determination of aquaporin Z (AqpZ) by ssNMR in <i>Escherichia coli</i> inner membranes. To enhance the signal sensitivity of AqpZ, we optimized protein overexpression and removed outer membrane components. To suppress the interference of background proteins, we used a "dual-media" expression approach and antibiotic treatment. Using 1017 distance restraints obtained from two-dimensional ¹³C-¹³C spectra based on the complete chemical shift assignments, the 1.7-Å ssNMR structure of AqpZ is determined in <i>E. coli</i> inner membranes. This cellular ssNMR structure determination paves the way for analyzing the atomic structural details for membrane proteins in native cellular membranes.