How to Prepare Membrane Proteins for Solid-State NMR: A Case Study on the α-Helical Integral Membrane Protein Diacylglycerol Kinase from E. coli

• Published in: Chembiochem : a European journal of chemical biology 2005-09, Vol.6 (9), p.1693-1700
• Main Authors: Lorch, Mark, Faham, Salem, Kaiser, Christoph, Weber, Ingrid, Mason, A. James, Bowie, James U, Glaubitz, Clemens
• Format: Article
• Language: English
• Subjects: Adenosine Triphosphate - pharmacology; biophysics; Crystallization; Diacylglycerol Kinase - chemistry; Diacylglycerol Kinase - metabolism; Escherichia coli; Escherichia coli - enzymology; Lipids - chemistry; membrane proteins; Membrane Proteins - chemistry; Membrane Proteins - metabolism; NMR spectroscopy; Nuclear Magnetic Resonance, Biomolecular - methods; Protein Structure, Secondary; protein structures
• ISSN: 1439-4227; 1439-7633; 1439-4227
• DOI: 10.1002/cbic.200500054

Several studies have demonstrated that it is viable to use microcrystalline preparations of water-soluble proteins as samples in solid-state NMR experiments.1-5 Here, we investigate whether this approach holds any potential for studying water-insoluble systems, namely membrane proteins. For this case study, we have prepared proteoliposomes and small crystals of the α-helical membrane-protein diacylglycerol kinase (DGK). Preparations were characterised by ¹³C- and ¹⁵N-cross-polarization magic-angle spinning (CPMAS) NMR. It was found that crystalline samples produce better-resolved spectra than proteoliposomes. This makes them more suitable for structural NMR experiments. However, reconstitution is the method of choice for biophysical studies by solid-state NMR. In addition, we discuss the identification of lipids bound to membrane-protein crystals by ³¹P-MAS NMR.