Theoretical Study On Mechanisms Of Cytochrome P450Enzyme-catalyzed Reactions Of Amines And Alkanes

Cytochrome P450enzymes are well-known as the most ubiquitous catalysts which essentially functionin both synthesis of endogenous compouds such as steroid, and metabolism of xenobiotics, such as drugs.Plenty of chemical reactions are catalyzed by P450s including hydroxylations of alkanes, oxidations ofalcohols, epoxidations of alkenes and arenes and dealkylations and deaminations of amines and itsalkane-substituted derivants.By means of density functional theory, we investigated the mechanisms of P450-catalyzeddeamination, inactivation and hydroxylation using cyclohexylamine, N-benzyl-N-cyclopropylamine andisopentane, which is an abbreviated model of cholesterol, as substrates. By calculating and analyzing theenergy barriers, geometries, spin densities and the corresponding kinetic isotope effects, not only severalknown experimental results are theoretically verified, but a few unknown significant points are proposed.For decades, the mechanism of the C-H bond oxidation which involved in the P450-catalyzeddeamination of cyclohexylamine has been in dispute. The calculated results reveal that such reaction is atypical two-state reactivity process due to the energy barriers for both quartet and doublet spin states areclose to each other. Moreover, the kinetic isotope effects of such process are comparable large, which is inaccord with the hydrogen atom transfer mechanism. Besides, the carbinolamine intermediate, which is theproduct of the C-H bond oxidation, could decompose to cyclopropanone and ammonia in a non-enzymeenvironment assisted by a water molecule.For N-benzyl-N-cyclopropylamine, the mechanism-based inactivator of P450, several proposedinactivation mechanisms are examined, the results initially rule out the single electron transfer mechanismfor its extremely high energy barriers, then by introducing the definition of proton-coupled electron transfermechanism as well as the variation of the spin density involving in the H-abtraction process, the validationof the hydrogen atom transfer mechanism is demonstrated. At the meantime, another reaction pathwaythrough which normal metabolites would formed is verified, and another new possible reaction route isfound. We believe that our novel theoretical findings would lay the foundation for the followingexperimental investigations.The two sequential hydroxylaions involved in the P450-catalyzed side-chain cleavage of cholesterol are theoretically studied. The results support the experimental observation that Cpd I is the major reactivespecies that catalyzes the hydroxyaltions, and such hydroxylations are indeed involves the hydrogen atomtransfer mechanism. Furthermore, focusing on the hydrogen bond formed between substrate and Cpd I, theanalyses of the energy barriers, geometries reveal several impacts that such hydrogen bond has on thereaction system. Finally, we perform a series of tests on the up-to-date dispersion-corrected densityfunctionals, and provide several valuable suggestions on how to opt for appropriate functionals duringtheoretical calculations.