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Characterization of Membrane Protein‐Lipid Interactions in Unfolded OmpX with Enhanced Time Resolution by Hyperpolarized NMR
Author(s) -
Kim Jihyun,
Mandal Ratnamala,
Hilty Christian
Publication year - 2020
Publication title -
chembiochem
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.05
H-Index - 126
eISSN - 1439-7633
pISSN - 1439-4227
DOI - 10.1002/cbic.202000271
Subject(s) - chemistry , micelle , nuclear magnetic resonance spectroscopy , phosphocholine , crystallography , membrane , urea , nmr spectra database , nuclear overhauser effect , guanidine , analytical chemistry (journal) , nuclear magnetic resonance , phospholipid , spectral line , phosphatidylcholine , stereochemistry , chromatography , organic chemistry , biochemistry , aqueous solution , physics , astronomy
Proton nuclear spins of dodecyl phosphocholine molecules below the critical micelle concentration are hyperpolarized by using dissolution dynamic nuclear polarization (D‐DNP). NMR signal enhancements of 1210±400 and 1610±550 are obtained at 9.4 T, for choline methyls in the head group of the lipid and for the tail‐end methyl group, respectively. This polarization is transferred to the unfolded protein through the nuclear Overhauser effect, after dilution to a final denaturant concentration of 0.8 M urea. As a result, the amide and aromatic side‐chain signals of the protein are increased up to sixfold. Selective inversion pulses applied either on the head‐group or tail‐group of the lipid are used to identify the source of the transferred polarization. The normalized cross‐relaxation rates of σ N,tail =−1.8±0.1 s −1 M −1 and σ N,head =−0.5±0.3 s −1 M −1 are obtained, showing a larger polarization transfer from the tail groups. These cross‐relaxation rates are determined at a low urea concentration, which constitutes refolding conditions for the protein. The sensitivity enhancement by D‐DNP permits to access these conditions with a measurement time on the order of seconds, and may further open the possibility to investigate structural changes in membrane proteins during folding.