Solution NMR studies of the integral membrane proteins OmpX and OmpA from Escherichia coli

• Published in: FEBS Letters Vol. 504; no. 3; pp. 173 - 178
• Main Authors: Fernández, César, Hilty, Christian, Bonjour, Sophie, Adeishvili, Koba, Pervushin, Konstantin, Wüthrich, Kurt
• Format: Book Review Journal Article
• Language: English
• Published: England Elsevier B.V 31.08.2001
• Subjects: 2D, two-dimensional; 3D, three-dimensional; Bacterial Outer Membrane Proteins - chemistry; Cell Membrane - chemistry; COSY, correlation spectroscopy; DHPC, dihexanoylphosphatidylcholine (1,2-dihexanoyl-sn-glycero-3-phosphocholine); DPC, dodecylphosphocholine; Escherichia coli - chemistry; Escherichia coli Proteins; Hydrolases; Isotope labeling; Magnetic Resonance Spectroscopy - methods; Membrane protein; Models, Molecular; NMR structure; NOE, nuclear Overhauser effect; NOESY, NOE spectroscopy; OmpA, outer membrane protein A from E. coli; OmpX, outer membrane protein X from E. coli; Outer membrane protein A from E. coli; Outer membrane protein X from E. coli; ppm, parts per million; Transverse relaxation-optimized spectroscopy; TROSY, transverse relaxation-optimized spectroscopy; TROSY, transverse relaxation-optimized spectroscopy; NMR structure; Membrane protein; OmpA, outer membrane protein A from E. coli; COSY, correlation spectroscopy; DHPC, dihexanoylphosphatidylcholine (1,2-dihexanoyl- sn-glycero-3-phosphocholine); NOE, nuclear Overhauser effect; 3D, three-dimensional; 2D, two-dimensional; Outer membrane protein X from E. coli; DPC, dodecylphosphocholine; NOESY, NOE spectroscopy; Outer membrane protein A from E. coli; Transverse relaxation-optimized spectroscopy; OmpX, outer membrane protein X from E. coli; ppm, parts per million; Isotope labeling
• ISSN: 0014-5793; 1873-3468
• DOI: 10.1016/S0014-5793(01)02742-9

Membrane proteins are usually solubilized in polar solvents by incorporation into micelles. Even for small membrane proteins these mixed micelles have rather large molecular masses, typically beyond 50 000 Da. The NMR technique TROSY (transverse relaxation-optimized spectroscopy) has been developed for studies of structures of this size in solution. In this paper, strategies for the use of TROSY-based NMR experiments with membrane proteins are discussed and illustrated with results obtained with the Escherichia coli integral membrane proteins OmpX and OmpA in mixed micelles with the detergent dihexanoylphosphatidylcholine (DHPC). For OmpX, complete sequence-specific NMR assignments have been obtained for the polypeptide backbone. The 13C chemical shifts and nuclear Overhauser effect data then resulted in the identification of the regular secondary structure elements of OmpX/DHPC in solution, and in the collection of an input of conformational constraints for the computation of the global fold of the protein. For OmpA, the NMR assignments are so far limited to about 80% of the polypeptide chain, indicating different dynamic properties of the reconstituted OmpA β-barrel from those of OmpX. Overall, the present data demonstrate that relaxation-optimized NMR techniques open novel avenues for studies of structure, function and dynamics of integral membrane proteins.