Probing the transmembrane structure and dynamics of microsomal NADPH-cytochrome P450 oxidoreductase by solid-state NMR.
NADPH-cytochrome P450 oxidoreductase (CYPOR) is an essential redox partner of the cytochrome P450 (cyt P450) superfamily of metabolic enzymes. In the endoplasmic reticulum of liver cells, such enzymes metabolize ~75% of the pharmaceuticals in use today. It is known that the transmembrane domain of CYPOR plays a crucial role in aiding the formation of a complex between CYPOR and cyt P450. Here we present the transmembrane structure, topology, and dynamics of the FMN binding domain of CYPOR in a native membrane-like environment. Our solid-state NMR results reveal that the N-terminal transmembrane domain of CYPOR adopts an α-helical conformation in the lipid membrane environment. Most notably, we also show that the transmembrane helix is tilted ~13° from the lipid bilayer normal, and exhibits motions on a submillisecond timescale including rotational diffusion of the whole helix and fluctuation of the helical director axis. The approaches and the information reported in this study would enable further investigations on the structure and dynamics of the full-length NADPH-cytochrome P450 oxidoreductase and its interaction with other membrane proteins in a membrane environment.
Biophysics and Department of Chemistry, University of Michigan, Ann Arbor, Michigan.
Rui
R
Huang
Biophysics and Department of Chemistry, University of Michigan, Ann Arbor, Michigan.
Kazutoshi
K
Yamamoto
Biophysics and Department of Chemistry, University of Michigan, Ann Arbor, Michigan.
Meng
M
Zhang
Biophysics and Department of Chemistry, University of Michigan, Ann Arbor, Michigan.
Nataliya
N
Popovych
National High Magnetic Field Lab, Tallahassee, Florida.
Ivan
I
Hung
Department of Anesthesiology, University of Michigan, Ann Arbor, Michigan; Veterans Administration Medical Center, Ann Arbor, Michigan.
Sang-Choul
SC
Im
National High Magnetic Field Lab, Tallahassee, Florida.
Zhehong
Z
Gan
Department of Anesthesiology, University of Michigan, Ann Arbor, Michigan; Veterans Administration Medical Center, Ann Arbor, Michigan.
Lucy
L
Waskell
Biophysics and Department of Chemistry, University of Michigan, Ann Arbor, Michigan. Electronic address: ramamoor@umich.edu.
Ayyalusamy
A
Ramamoorthy
D000818:Animals; D002462:Cell Membrane / Q000378:metabolism*; D005486:Flavin Mononucleotide / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy; D008565:Membrane Proteins / Q000378:metabolism; D008861:Microsomes / Q000201:enzymology*; D008958:Models, Molecular; D009251:NADPH-Ferrihemoprotein Reductase / Q000737:chemistry* / Q000378:metabolism*; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D051381:Rats
D008565:Membrane Proteins; D005486:Flavin Mononucleotide; D009251:NADPH-Ferrihemoprotein Reductase