Optimizing the alpha(1B)-adrenergic receptor for solution NMR studies
Matthias Schuster, Mattia Deluigi, Milica Pantic, Santiago Vacca, Christian Baumann, Daniel J Scott, Andreas Pluckthun, Oliver Zerbe
BIOCHIMICA ET BIOPHYSICA ACTA-BIOMEMBRANES | ELSEVIER | Published : 2020
Sample preparation for NMR studies of G protein-coupled receptors faces special requirements: Proteins need to be stable for prolonged measurements at elevated temperatures, they should ideally be uniformly labeled with the stable isotopes 13C, 15N, and all carbon-bound protons should be replaced by deuterons. In addition, certain NMR experiments require protonated methyl groups in the presence of a perdeuterated background. All these requirements are most easily satisfied when using Escherichia coli as the expression host. Here we describe a workflow, starting from a temperature-stabilized mutant of the α1B-adrenergic receptor, obtained using the CHESS methodology, into an even more stable ..View full abstract
Awarded by Swiss National Science Foundation
Awarded by Research Foundation of the University of Zurich
This project is supported by a grant from the Swiss National Science Foundation (grant no. 310030_179314 to O.Z.) and from the Research Foundation of the University of Zurich (grant no. FK-18-083 to C.B.). We deeply thank Franz Hagn for a gift of the MSP Delta H5 expression plasmid.