Abstract
This work investigates the Y326I point mutation effect on the kinetics of oxidative deamination of phenylethylamine (PEA) catalyzed by the monoamine oxidase B (MAO B) enzyme. PEA is a neuromodulator capable of affecting the plasticity of the brain and is responsible for the mood enhancing effect caused by physical exercise. Due to a similar functionality, PEA is often regarded as an endogenous amphetamine. The rate limiting step of the deamination was simulated at the multiscale level, employing the Empirical Valence Bond approach for the quantum treatment of the involved valence states, whereas the environment (solvated protein) was represented with a classical force field. A comparison of the reaction free energy profiles delivered by simulation of the reaction in the wild type MAO B and its Y326I mutant yields an increase in the barrier by 1.06 kcal mol-1 upon mutation, corresponding to a roughly 6-fold decrease in the reaction rate. This is in excellent agreement with the experimental kinetic studies. Inspection of simulation trajectories reveals possible sources of the point mutation effect, namely vanishing favorable electrostatic interactions between PEA and a Tyr326 side chain and an increased amount of water molecules at the active site due to the replacement of tyrosine by a less spacious isoleucine residue, thereby increasing the dielectric shielding of the catalytic environment provided by the enzyme.
Highlights
monoamine oxidase enzymes (MAOs) enzymes exist in two forms, A and B (MAO A, monoamine oxidase B (MAO B)), which share about 70% of the sequence identity,[10] but differ in substrate specificity.[2,11] both MAO A and MAO B share a wide array of common substrates
It has been accepted that the catalytic step of amine oxidation includes the cleavage of a C–H bond at the methylene group vicinal to the amino group, i.e., the a-C–H bond, accompanied by the hydrogen transfer to the flavin adenine dinucleotide (FAD) prosthetic group covalently bound to the cysteine residue.[12]
MAO B was used as reference, simulation in the wild type (WT) enzyme by definition reproduces the experimental barrier of 17.29 kcal molÀ1; the change in the barrier when passing from WT to the mutated enzyme was obtained from simulation of the reaction in
Summary
MAO enzymes exist in two forms, A and B (MAO A, MAO B), which share about 70% of the sequence identity,[10] but differ in substrate specificity.[2,11] both MAO A and MAO B share a wide array of common substrates. We recently demonstrated that the reaction rate for the protonated adrenaline substrate is extremely low.[64] In order to make the computed barrier fully comparable with experimental data, the barrier should be corrected for the free energy of deprotonation of PEA, because the experimental measurement includes this correction by definition This requires that the pKa value of PEA be estimated within the active site of MAO B, which is all but a trivial task.[65] the assumption that the pKa value of PEA does not change much upon mutation appears to be reasonable, the deprotonation corrections are canceled out in the present treatment. The results contribute to an improved understanding of the chemical processes involved in the metabolism of neurotransmitters
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