title: Nanoscale electron transport measurements of immobilized cytochrome P450 proteins.
abstract: Gold nanopillars, functionalized with an organic self-assembled monolayer, can be used to measure the electrical conductance properties of immobilized proteins without aggregation. Measurements of the conductance of nanopillars with cytochrome P450 2C9 (CYP2C9) proteins using conducting probe atomic force microscopy demonstrate that a correlation exists between the energy barrier height between hopping sites and CYP2C9 metabolic activity. Measurements performed as a function of tip force indicate that, when subjected to a large force, the protein is more stable in the presence of a substrate. This agrees with the hypothesis that substrate entry into the active site helps to stabilize the enzyme. The relative distance between hopping sites also increases with increasing force, possibly because protein functional groups responsible for electron transport (ETp) depend on the structure of the protein. The inhibitor sulfaphenazole, in addition to the previously studied aniline, increased the barrier height for electron transfer and thereby makes CYP2C9 reduction more difficult and inhibits metabolism. This suggests that P450 Type II binders may decrease the ease of ETp processes in the enzyme, in addition to occupying the active site.
authors: Department of Pharmaceutical Sciences, West Virginia University, Morgantown, WV 26506-9530, USA.
authors: Christopher D
authors: CD
authors: Bostick
authors: Darcy R
authors: DR
authors: Flora
authors: Peter M
authors: PM
authors: Gannett
authors: Timothy S
authors: TS
authors: Tracy
authors: David
authors: D
authors: Lederman
mesh_terms: D000814:Aniline Compounds / Q000737:chemistry*; D020134:Catalytic Domain; D065729:Cytochrome P-450 CYP2C9 / Q000378:metabolism; D003577:Cytochrome P-450 Enzyme System / Q000737:chemistry*; D003622:Dapsone / Q000737:chemistry; D004553:Electric Conductivity; D004579:Electron Transport; D004583:Electrons; D005480:Flurbiprofen / Q000737:chemistry; D006046:Gold / Q000737:chemistry; D006801:Humans; D055767:Immobilized Proteins / Q000737:chemistry*; D053768:Metal Nanoparticles / Q000737:chemistry; D008855:Microscopy, Electron, Scanning; D011485:Protein Binding; D011487:Protein Conformation; D015202:Protein Engineering / Q000379:methods; D012825:Silicon / Q000737:chemistry; D013426:Sulfaphenazole / Q000737:chemistry
chemical_list: D000814:Aniline Compounds; D055767:Immobilized Proteins; D013426:Sulfaphenazole; D005480:Flurbiprofen; D006046:Gold; D003622:Dapsone; D003577:Cytochrome P-450 Enzyme System; C450260:CYP2C9 protein, human; D065729:Cytochrome P-450 CYP2C9; C023650:aniline; D012825:Silicon