Quadruple-resonance magic-angle spinning NMR spectroscopy of deuterated solid proteins.
(1)H-detected magic-angle spinning NMR experiments facilitate structural biology of solid proteins, which requires using deuterated proteins. However, often amide protons cannot be back-exchanged sufficiently, because of a possible lack of solvent exposure. For such systems, using (2)H excitation instead of (1)H excitation can be beneficial because of the larger abundance and shorter longitudinal relaxation time, T1, of deuterium. A new structure determination approach, "quadruple-resonance NMR spectroscopy", is presented which relies on an efficient (2)H-excitation and (2)H-(13)C cross-polarization (CP) step, combined with (1)H detection. We show that by using (2)H-excited experiments better sensitivity is possible on an SH3 sample recrystallized from 30 % H2O. For a membrane protein, the ABC transporter ArtMP in native lipid bilayers, different sets of signals can be observed from different initial polarization pathways, which can be evaluated further to extract structural properties.
Leibniz Institute for Molecular Pharmacology, Robert Roessle Str. 10, 13125 Berlin (Germany). akbey@fmp-berlin.de.
Ümit
Ü
Akbey
Andrew J
AJ
Nieuwkoop
Sebastian
S
Wegner
Anja
A
Voreck
Britta
B
Kunert
Priyanga
P
Bandara
Frank
F
Engelke
Niels Chr
NC
Nielsen
Hartmut
H
Oschkinat
D001426:Bacterial Proteins / Q000737:chemistry*; D003903:Deuterium / Q000032:analysis; D001411:Geobacillus stearothermophilus / Q000737:chemistry*; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods*; D011487:Protein Conformation
D001426:Bacterial Proteins; D003903:Deuterium
deuteration; high sensitivity; proteins; quadruple-resonance MAS NMR spectroscopy; structure elucidation