Lipid-protein nanodiscs: possible application in high-resolution NMR investigations of membrane proteins and membrane-active peptides.
High-resolution NMR is shown to be applicable for investigation of membrane proteins and membrane-active peptides embedded into lipid-protein nanodiscs (LPNs). (15)N-Labeled K+-channel from Streptomyces lividans (KcsA) and the antibiotic antiamoebin I from Emericellopsis minima (Aam-I) were embedded in LPNs of different lipid composition. Formation of stable complexes undergoing isotropic motion in solution was confirmed by size-exclusion chromatography and (31)P-NMR spectroscopy. The 2D 1H-(15)N-correlation spectra were recorded for KcsA in the complex with LPN containing DMPC and for Aam-I in LPNs based on DOPG, DLPC, DMPC, and POPC. The spectra recorded were compared with those in detergent-containing micelles and small bicelles commonly used in high-resolution NMR spectroscopy of membrane proteins. The spectra recorded in LPN environments demonstrated similar signal dispersion but significantly increased (1)H(N) line width. The spectra of Aam-I embedded in LPNs containing phosphatidylcholine showed significant selective line broadening, thus suggesting exchange process(es) between several membrane-bound states of the peptide. (15)N relaxation rates were measured to obtain the effective rotational correlation time of the Aam-I molecule. The obtained value (approximately 40 nsec at 45 degrees C) is indicative of additional peptide motions within the Aam-I/LPN complex.
Shenkarev
Z O
ZO
Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow, 117997, Russia. zh@nmr.ru
Lyukmanova
E N
EN
Solozhenkin
O I
OI
Gagnidze
I E
IE
Nekrasova
O V
OV
Chupin
V V
VV
Tagaev
A A
AA
Yakimenko
Z A
ZA
Ovchinnikova
T V
TV
Kirpichnikov
M P
MP
Arseniev
A S
AS
D001426:Bacterial Proteins / Q000737:chemistry*; D006999:Hypocreales / Q000737:chemistry; D008055:Lipids / Q000737:chemistry*; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D008565:Membrane Proteins / Q000737:chemistry*; D049329:Nanostructures / Q000737:chemistry*; D054848:Peptaibols; D010455:Peptides / Q000737:chemistry*; D015221:Potassium Channels / Q000737:chemistry*
D001426:Bacterial Proteins; D008055:Lipids; D008565:Membrane Proteins; D054848:Peptaibols; D010455:Peptides; D015221:Potassium Channels; C107790:prokaryotic potassium channel; C006596:antiamoebin