Computational Insight into the Mechanism of the Irreversible Inhibition of Monoamine Oxidase Enzymes by the Antiparkinsonian Propargylamine Inhibitors Rasagiline and Selegiline

Abstract

Monoamine oxidases (MAO) are FAD-containing flavoenzymes that catalyze the degradation of a range of brain neurotransmitters, whose imbalance is extensively linked with the pathology of various neurological disorders. This is why MAOs have been the central pharmacological targets in treating neurodegeneration for more than 60 years. Still, despite this practical importance, the precise chemical mechanisms underlying the irreversible inhibition of the MAO B isoform with clinical drugs rasagiline (RAS) and selegiline (SEL) remained unknown. Here we employed a combination of MD simulations, MM-GBSA binding free energy evaluations, and QM cluster calculations to show the MAO inactivation proceeds in three steps, where, in the rate-limiting first step, FAD utilizes its N5 atom to abstracts a hydride anion from the inhibitor α-CH 2 group to ultimately give the final inhibitor-FAD adduct matching crystallographic data. The obtained free energy profiles reveal a lower activation energy for SEL by 1.2 kcal mol -1 and a higher reaction exergonicity by 0.8 kcal mol -1 , the former in excellent agreement with experimental ΔΔG ‡ EXP = 1.7 kcal mol -1 , thus rationalizing its higher in vivo reactivity over RAS. The calculated ΔG BIND energies confirm SEL binds better due to its bigger size and flexibility allowing it to optimize hydrophobic C-H•••π and π•••π interactions with residues throughout both of enzyme's cavities, particularly with FAD, Gln206 and four active site tyrosines, thus overcoming a larger ability of RAS to form hydrogen bonds that only position it in less reactive orientations for the hydride abstraction. Offered results elucidate structural determinants affecting the affinity and rates of the inhibition reaction that should be considered to co-operate when designing more effective compounds devoid of untoward effects, which are of utmost significance and urgency with the growing prevalence of brain diseases.

Hydride Abstraction as the Rate-Limiting Step of the Irreversible Inhibition of Monoamine Oxidase B by Rasagiline and Selegiline: A Computational Empirical Valence Bond Study

“…More importantly, this was followed by the quantum mechanics (QM) cluster calculations [ 54 ] at the DFT level, which revealed that the MAO inactivation proceeded through a three-step reaction, where, in the rate-limiting first step, the MAO enzyme used the FAD’s N5 atom to abstract a hydride anion from the inhibitor’s α-CH 2 group ( Figure 2 ), being in a full analogy with the MAO catalytic mechanism [ 55 , 56 , 57 , 58 ], thus confirming that both SEL and RAS are mechanism-based inhibitors. The complete reaction profile led from the bound inhibitors to the corresponding N5(FAD)-adducts, the latter being in excellent agreement with the crystallographic data on the inhibited enzyme, and further confirmed a better reactivity of SEL through both its lower activation barrier and higher overall exergonicity [ 52 ]. Still, although the obtained relative difference in the activation free energies among inhibitors of ΔΔ G ‡ = 1.2 kcal mol −1 was found to be in excellent agreement with the measured k inact values, which predicts a difference of ΔΔ G ‡ EXP = 1.7 kcal mol −1 , the absolute values of Δ G ‡ (SEL) = 27.9 kcal mol −1 and Δ G ‡ (RAS) = 29.1 kcal mol −1 clearly exceeded those predicted experimentally, although these were significantly lower than those calculated for several alternative mechanistic scenarios.…”

“…While we were not able to locate the corresponding literature data for rasagiline, we can assume that its p K a value should be roughly the same. Our earlier mechanistic proposal required unionized inhibitors to undergo the MAO inhibition [ 52 ], since it is significantly easier to detach the hydride anion from the neutral system and generate the carbocation, than it is from the already monocationic system that is transformed into dicationic compounds upon the hydride abstraction. This is justified considering the mentioned p K a value for selegiline being close to the physiological pH value, but also knowing the hydrophobic nature of the MAO active site, which was clearly demonstrated experimentally [ 5 , 34 ] and confirmed computationally [ 69 ].…”

“…In our recent work [ 52 ], we performed molecular dynamics simulations [ 53 ] to investigate the binding of SEL and RAS within the MAO B active site and identified residues predominantly responsible for the successful binding. In addition, the calculated MM-GBSA binding free-energies effectively reproduced a trend in the measured K i and IC 50 data, and thus confirmed that SEL binds better due to its bigger size and flexibility, allowing it to optimize hydrophobic C–H∙∙∙π and π∙∙∙π interactions with residues throughout both of the enzyme’s cavities, particularly with FAD, Gln206 and four active site tyrosines (Tyr435, Tyr398, Tyr60, Tyr326).…”

Relevance of Hydrogen Bonds for the Histamine H2 Receptor-Ligand Interactions: A Lesson from Deuteration

“…The effect of the rest of the protein environment was considered with the CPCM implicit solvation model using a dielectric constant of ε = 4.0, as suggested by Himo and co-workers [28], and a dielectric constant of ε = 78.4 for the aqueous solution, all in line with our previous reports [13], where we also demonstrated that a potential increase in the former dielectric constant to ε = 20.0 lowers the accuracy of the obtained results and even predicts wrong trends among ligands. Additionally, in our experience, such a truncated cluster-continuum model of the entire protein turned very useful in rationalizing various aspects of the catalytic activity [29], selectivity [30] and inhibition [31] of the monoamine oxidase family of enzymes, and is broadly used by different groups to decipher various biological phenomena [32][33][34][35][36], which justifies its use here. We note in passing that, despite its practical usefulness, the proposed value of ε = 4 is, in some cases, apparently too small to prevent the proton transfer from protonated to anionic residues, which then occurred spontaneously during the geometry optimization, provided the involved pair is in close vicinity.…”

Novel Harmicines with Improved Potency against Plasmodium

“…The binding free energies, Δ G BIND , of each ligand within the Pf Hsp90 ATP binding site were calculated using the established MM-GBSA protocol [ 35 , 36 ] available in AmberTools16 [ 32 ], and in line with our earlier reports [ 15 , 37 , 38 ]. MM-GBSA is a widely used method for binding free energy calculations from snapshots of MD trajectory with an estimated standard error of 1–3 kcal mol −1 [ 36 ].…”