Observation of ligand binding to cytochrome P450 BM-3 by means of solid-state NMR spectroscopy.
A broad understanding of the binding modes of ligands and inhibitors to cytochrome P450 is vital for the development of new drugs. We investigated ligand binding in a site-specific fashion on cytochrome P450 BM-3 from Bacillus megaterium, a 119 kDa paramagnetic enzyme, using solid-state magic angle spinning nuclear magnetic resonance methods. Selective labeling and longitudinal relaxation effects were utilized to identify the peaks in a site-specific fashion and to provide evidence for binding. Well-resolved one-dimensional and two-dimensional NMR spectra of cytochrome P450 BM-3 reveal shifts upon binding of its substrate, N-palmitoylglycine. These data are consistent with the crystallographic result that a biochemically important amino acid residue, Phe87, moves upon ligation. This experimental scheme provides a tool for probing ligand binding for complex systems.
Jovanovic
Tijana
T
Department of Chemistry, Columbia University, 3000 Broadway, New York, New York 10027, USA.
McDermott
Ann E
AE
D001410:Bacillus megaterium / Q000201:enzymology; D002247:Carbon Isotopes; D003577:Cytochrome P-450 Enzyme System / Q000737:chemistry*; D008024:Ligands; D009682:Magnetic Resonance Spectroscopy / Q000379:methods* / Q000592:standards; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D012015:Reference Standards
D002247:Carbon Isotopes; D008024:Ligands; D003577:Cytochrome P-450 Enzyme System