Quantum mechanics/molecular mechanics modeling of drug metabolism:Mexiletine N-hydroxylation by cytochrome P450 1A2

The mechanism of cytochrome P450­(CYP)-catalyzed hydroxylation of primary amines is currently unclear and is relevant to drug metabolism; previous small model calculations have suggested two possible mechanisms: direct N-oxidation and H-abstraction/rebound. We have modeled the N-hydroxylation of (<i>R</i>)-mexiletine in CYP1A2 with hybrid quantum mechanics/molecular mechanics (QM/MM) methods, providing a more detailed and realistic model. Multiple reaction barriers have been calculated at the QM­(B3LYP-D)/MM­(CHARMM27) level for the direct N-oxidation and H-abstraction/rebound mechanisms. Our calculated barriers indicate that the direct N-oxidation mechanism is preferred and proceeds via the doublet spin state of Compound I. Molecular dynamics simulations indicate that the presence of an ordered water molecule in the active site assists in the binding of mexiletine in the active site, but this is not a prerequisite for reaction via either mechanism. Several active site residues play a role in the binding of mexiletine in the active site, including Thr124 and Phe226. This work reveals key details of the N-hydroxylation of mexiletine and further demonstrates that mechanistic studies using QM/MM methods are useful for understanding drug metabolism

Theoretical Studies of Cytochrome P450

The cytochromes P450 are a large enzyme family that is found in all living organisms and takes part in both endogenous and exogenous metabolism. They are primarily mono-oxygenases and perform a wide range of reactions. In contrast to many other enzymes, they have been optimised to react with a wide range of substrates instead of having high reaction rates. In the active site, these enzymes have a haem group, a porphyrin ring with an iron ion in the centre, and a cysteine amino acid bound to the iron. During the reaction cycle, this complex is turned into a highly oxidative complex, which enables the enzyme to oxidise even aliphatic carbons. We study various aspects of the functionality of these enzymes and other related haem-containing proteins with theoretical methods. For example, we discuss how nature has tuned the functionality of different haem proteins by using different axial ligands, and go into depth on the elusive properties of a mobile proton in haem peroxidases. Furthermore, we study the transition state in aliphatic hydroxylation, nitrogen oxidation, sulphur oxidation, and sulphoxide oxidation in cytochrome P450 with density functional theory and suggest a qualitative model to predict the activation energy of the aliphatic hydroxylation reaction. By constructing a transition-state force field and combining it with the qualitative model, we study the hydroxylation reactivity of two drugs in two cytochrome P450 isoforms. Finally, using molecular dynamics, we provide the first calculation of the free energy for moving a substrate molecule from bulk water solution into the active site of a cytochrome P450, and we also study the dynamics of water molecules in the active-site cavity

Theoretical Study of the Cytochrome P450 Mediated Metabolism of Phosphorodithioate Pesticides

The toxicity of phosphorodithioate pesticides is due to the formation of the active oxane product through desulfurization by cytochrome P450 enzymes, both in humans and insects. During this desulfurization, inhibition of cytochrome P450 and a loss of heme has been observed. Here, we study the mechanism of desulfurization and inhibition with density functional theory, using the B3LYP functional with and without dispersion correction. The results show that a reaction mechanism initiated by sulfur oxidation is most likely, with a reaction barrier of 47 kJ/mol. The sulfur oxidation is followed by a ring-closing mechanism with a barrier of 28 kJ/mol relative to the sulfur-oxidized intermediate. The enzymatic contribution to the ring-closing is very small. It is also shown that the apparent loss of heme might be due to the formation of a previously unknown inhibition complex, which changes the aromatic conjugation of the porphyrin ring. We also show that including dispersion correction has significant effects on a ring closure transition state (∼30 kJ/mol), whereas effects on the other steps in the reaction are relatively small (4–15 kJ/mol)

WhichCyp SVM modelling data

<p>output from R</p

p450 metabolism examples

<p>two examples of P450 mediated drug metabolism</p

phase1metabolismfigure

<p>figure of enzymes contributing to phase I metabolism</p