Solution-state NMR spectroscopy of membrane proteins in detergent micelles: structure of the Klebsiella pneumoniae outer membrane protein A, KpOmpA.
Structure determination of integral membrane proteins is one of the most important challenges of structural biology. Over the last 7 years, solution-state NMR spectroscopy has become an increasingly useful approach for 3D structure determination and dynamical analysis of membrane proteins solubilized in detergent micelles. We describe herein an ensemble of methods applied in this context, including isotopic labelling, in vitro refolding procedure, and state-of-the-art NMR experiments required for the structure determination of high molecular weight molecular complexes. Furthermore, the basic principles of spectrum interpretation and 3D structure calculation are reported. This approach is illustrated with the structural study of the transmembrane domain of the outer membrane protein A from Klebsiella pneumoniae (KpOmpA).
Renault
Marie
M
UPS, Institut de Pharmacologie et de Biologie Structurale, Université de Toulouse, CNRS, UMR 5089, Toulouse, France.
Saurel
Olivier
O
Demange
Pascal
P
Reat
Valérie
V
Milon
Alain
A
D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry* / Q000378:metabolism; D003902:Detergents / Q000737:chemistry*; D007711:Klebsiella pneumoniae / Q000378:metabolism*; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D008565:Membrane Proteins / Q000737:chemistry*; D008823:Micelles
D001425:Bacterial Outer Membrane Proteins; D003902:Detergents; D008565:Membrane Proteins; D008823:Micelles