17528-90-4Relevant articles and documents
A Modified Arrhenius Approach to Thermodynamically Study Regioselectivity in Cytochrome P450-Catalyzed Substrate Conversion
Luirink, Rosa A.,Verkade-Vreeker, Marlies C. A.,Commandeur, Jan N. M.,Geerke, Daan P.
, p. 1461 - 1472 (2020/03/03)
The regio- (and stereo-)selectivity and specific activity of cytochrome P450s are determined by the accessibility of potential sites of metabolism (SOMs) of the bound substrate relative to the heme, and the activation barrier of the regioselective oxidation reaction(s). The accessibility of potential SOMs depends on the relative binding free energy (ΔΔGbind) of the catalytically active substrate-binding poses, and the probability of the substrate to adopt a transition-state geometry. An established experimental method to measure activation energies of enzymatic reactions is the analysis of reaction rate constants at different temperatures and the construction of Arrhenius plots. This is a challenge for multistep P450-catalyzed processes that involve redox partners. We introduce a modified Arrhenius approach to overcome the limitations in studying P450 selectivity, which can be applied in multiproduct enzyme catalysis. Our approach gives combined information on relative activation energies, ΔΔGbind values, and collision entropies, yielding direct insight into the basis of selectivity in substrate conversion.
Cytochrome P450 metabolic activities in the small intestine of cynomolgus macaques bred in cambodia, china, and indonesia
Nakanishi, Yasuharu,Yamashita, Hiroyuki,Yoshikawa, Tsuyoshi,Tominaga, Takeshi,Nojiri, Koichiro,Sunaga, Yoshiharu,Muneoka, Atsunobu,Iwasaki, Kazuhide,Utoh, Masahiro,Nakamura, Chika,Yamazaki, Hiroshi,Uno, Yasuhiro
, p. 510 - 513 (2014/02/14)
Summary: Cynomolgus macaques, used in drug metabolism studies due to their evolutionary closeness to humans, are mainly bred in Asian countries, including Cambodia, China, and Indonesia. Cytochromes P450 (P450s) are important drug-metabolizing enzymes, present in the liver and small intestine, major drug metabolizing organs. Previously, our investigation did not find statistically significant differences in hepatic P450 metabolic activities measured in cynomolgus macaques bred in Cambodia (MacfaCAM) and China (MacfaCHN). In the present study, P450 metabolic activity was investigated in the small intestine of MacfaCAM and MacfaCHN, and cynomolgus macaques bred in Indonesia (MacfaIDN) using P450 substrates, including 7-ethoxyresorufin, coumarin, bupropion, paclitaxel, diclofenac, S-mephenytoin, bufuralol, chlorzoxazone, and testosterone. The results indicated that P450 metabolic activity of the small intestine was not statistically significantly different (2.0-fold) in MacfaCAM, MacfaCHN, and MacfaIDN. In addition, statistically significant sex differences were not observed (2.0-fold) in any P450 metabolic activity in MacfaCAM as supported by mRNA expression results. These results suggest that P450 metabolic activity of the small intestine does not significantly differ statistically among MacfaCAM, MacfaCHN, and MacfaIDN.
Regio- and stereoselectivity of P450-catalysed hydroxylation of steroids controlled by laboratory evolution
Kille, Sabrina,Zilly, Felipe E.,Acevedo, Juan P.,Reetz, Manfred T.
scheme or table, p. 738 - 743 (2012/02/15)
A current challenge in synthetic organic chemistry is the development of methods that allow the regio- and stereoselective oxidative C - H activation of natural or synthetic compounds with formation of the corresponding alcohols. Cytochrome P450 enzymes enable C - H activation at non-activated positions, but the simultaneous control of both regio- and stereoselectivity is problematic. Here, we demonstrate that directed evolution using iterative saturation mutagenesis provides a means to solve synthetic problems of this kind. Using P450 BM3(F87A) as the starting enzyme and testosterone as the substrate, which results in a 1:1 mixture of the 2β- and 15β-alcohols, mutants were obtained that are 96 - 97% selective for either of the two regioisomers, each with complete diastereoselectivity. The mutants can be used for selective oxidative hydroxylation of other steroids without performing additional mutagenesis experiments. Molecular dynamics simulations and docking experiments shed light on the origin of regio- and stereoselectivity.
