1,2,3-Triazole-heme interactions in cytochrome P450: functionally competent triazole-water-heme complexes.
In comparison to imidazole (IMZ) and 1,2,4-triazole (1,2,4-TRZ), the isosteric 1,2,3-triazole (1,2,3-TRZ) is unrepresented among cytochrome P450 (CYP) inhibitors. This is surprising because 1,2,3-TRZs are easily obtained via "click" chemistry. To understand this underrepresentation of 1,2,3-TRZs among CYP inhibitors, thermodynamic and density functional theory computational studies were performed with unsubstituted IMZ, 1,2,4-TRZ, and 1,2,3-TRZ. The results indicate that the lower affinity of 1,2,3-TRZ for the heme iron includes a large unfavorable entropy term likely originating in solvent-1,2,3-TRZ interactions; the difference is not solely due to differences in the enthalpy of heme-ligand interactions. In addition, the 1,2,3-TRZ fragment was incorporated into a well-established CYP3A4 substrate and mechanism-based inactivator, 17-α-ethynylestradiol (17EE), via click chemistry. This derivative, 17-click, yielded optical spectra consistent with low-spin ferric heme iron (type II) in contrast to 17EE, which yields a high-spin complex (type I). Furthermore, the rate of CYP3A4-mediated metabolism of 17-click was comparable to that of 17EE, with a different regioselectivity. Surprisingly, continuous-wave electron paramagnetic resonance (EPR) and HYSCORE EPR spectroscopy indicate that 17-click does not displace water from the sixth axial ligand position of CYP3A4 as expected for a type II ligand. We propose a binding model in which 17-click pendant 1,2,3-TRZ hydrogen bonds with the sixth axial water ligand. The results demonstrate the potential for 1,2,3-TRZ to form metabolically labile water-bridged low-spin heme complexes, consistent with recent evidence that nitrogenous type II ligands of CYPs can be efficiently metabolized. The specific case of [CYP3A4·17-click] highlights the risk of interpreting CYP-ligand complex structure on the basis of optical spectra.
Department of Medicinal Chemistry, Box 357610, University of Washington, Seattle, WA 98195-7610, USA.
Kip P
KP
Conner
Preethi
P
Vennam
Caleb M
CM
Woods
Matthew D
MD
Krzyaniak
Michael K
MK
Bowman
William M
WM
Atkins
D057930:Click Chemistry; D051544:Cytochrome P-450 CYP3A / Q000737:chemistry; D065692:Cytochrome P-450 CYP3A Inhibitors; D065607:Cytochrome P-450 Enzyme Inhibitors; D003577:Cytochrome P-450 Enzyme System / Q000737:chemistry*; D004578:Electron Spin Resonance Spectroscopy; D004997:Ethinyl Estradiol / Q000031:analogs & derivatives / Q000138:chemical synthesis / Q000737:chemistry; D006418:Heme / Q000737:chemistry*; D007093:Imidazoles / Q000737:chemistry; D007536:Isomerism; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D011485:Protein Binding; D011789:Quantum Theory; D013816:Thermodynamics; D014230:Triazoles / Q000138:chemical synthesis / Q000737:chemistry*; D014867:Water / Q000737:chemistry*
D065692:Cytochrome P-450 CYP3A Inhibitors; D065607:Cytochrome P-450 Enzyme Inhibitors; D007093:Imidazoles; D014230:Triazoles; D014867:Water; D004997:Ethinyl Estradiol; D006418:Heme; C029899:imidazole; D003577:Cytochrome P-450 Enzyme System; D051544:Cytochrome P-450 CYP3A