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 Table of Contents  
Year : 2019  |  Volume : 17  |  Issue : 4  |  Page : 354-360

Molecular docking study of binding modes of amphetamine, cathine, and cathinone to monoamine oxidase B

Department of MLT, College of Applied Medical Sciences, Jazan University, Jazan, Kingdom of Saudi Arabia

Date of Submission30-Apr-2019
Date of Decision26-Sep-2019
Date of Acceptance03-Nov-2019
Date of Web Publication14-Feb-2020

Correspondence Address:
Jerah A Ahmed
The University Street, Building No. 6461, Jazan, Kingdom of Saudi Arabia
Kingdom of Saudi Arabia
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Source of Support: None, Conflict of Interest: None

DOI: 10.4103/AZMJ.AZMJ_75_19

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Background Molecular docking is commonly used in the study of small-molecule–protein interaction. Amphetamine (AMPH), cathine (CATHI), and cathinone (CATHO) bind in silico with monoamine oxidase B (MAO B) with relatively low affinity.
Objective The purpose of the study was to determine the binding modes of AMPH, CATHI, and CATHO with the enzyme MAO B. Molecular docking software, AutoDock 4.2, was used to study the interaction of these ligands with MAO B.
Materials and methods Molecular docking was performed using the molecular docking software AutoDock, version 4.2. graphic user interface, AutoDock Tools, was used to prepare protein and ligands for docking.
Results The estimated ΔG of binding (free energy of binding) was −6.25 kcal/mol for AMPH, −6.05 kcal/mol for CATHI, and −6.24 kcal/mol for CATHO. Respective Ki (inhibitor constant) values were 2.61×10−5 M, 3.69×10−5 M, and 2.69×10−5 M. All three ligands show similar interactions within active site which include hydrophobic and hydrogen-bonding interactions. When compared with binding affinity of known inhibitors of MAO, safinamide, the binding affinity of these inhibitors is substantially less. But, they bind in the same active site and share some common interactions with active site residues. Similarity in binding modes of AMPH, CATHI, and CATHO is due to their similar structures.
Conclusion It can be speculated that nervous stimulant activity of AMPH, CATHI, and CATHO could be due to inhibition of MAO. This enzyme catalyzes the oxidative deamination of monoamine neurotransmitters such as serotonin and dopamine and reduces the level of these neurotransmitters. Inhibition of this enzyme might result in more sustained levels of these neurotransmitters. Our docking study shows that AMPH, CATHI, and CATHO inhibit MAO B with free energy ranging from −6.05 to −6.25 kcal/mol and Ki ranging from 2.61×10−5 to 3.69×10−5 M.

Keywords: amphetamine, cathine, cathinone, khat, molecular docking, monoamine oxidase B, neurotransmitter

How to cite this article:
Ahmed JA. Molecular docking study of binding modes of amphetamine, cathine, and cathinone to monoamine oxidase B. Al-Azhar Assiut Med J 2019;17:354-60

How to cite this URL:
Ahmed JA. Molecular docking study of binding modes of amphetamine, cathine, and cathinone to monoamine oxidase B. Al-Azhar Assiut Med J [serial online] 2019 [cited 2020 Sep 29];17:354-60. Available from: http://www.azmj.eg.net/text.asp?2019/17/4/354/278400

  Introduction Top

Khat is chewed mainly for its intoxicating euphoric effect which is attributed to its active ingredients, cathine (CATHI), and cathinone (CATHO). These are nervous stimulants that exhibit an effect similar to that of amphetamine (AMPH). CATHI and CATHO are phenylalkylamines found in khat that have structural similarity to AMPH and noradrenaline [1]. CATHO, the main active alkaloid of khat, has AMPH −like properties [2],[3] and like AMPH it is considered as an indirect dopaminergic agonistic drug [4],[5]. In addition, (−)-CATHO also releases serotonin from its striatal stores, an action similar to (+)-AMPH [2]. The effect of (−)-CATHO on neurotransmission is similar to that of (+)-AMPH although at 2–10 times lesser potency. In terms of potency, khat alkaloids lie between caffeine and AMPH [6]. The central nervous stimulant potency of CATHO is about half of AMPH [7]. CATHI (norpseudoephedrine) and norephedrine are two other pharmacologically active compounds in khat which are less potent stimulants [8]. Their effect on the nervous system is also qualitatively similar to that of AMPH [9],[10],[11].

Monoamine oxidase [MAO, EC; amine : oxygen oxidoreductase (deaminating, flavin-containing)] is an flavin adenine dinucleotide (FAD)-containing enzyme. It is found in neuronal, glial, and other cells and is located in the mitochondrial outer membrane of these cells. It is involved in the metabolism of neurotransmitter amines like dopamine, serotonin, phenylethylamine, and norepinephrine. In the metabolism of these monoamine neurotransmitters, the enzyme catalyzes oxidative deamination of monoamine. It is thought to play an important role in many psychiatric and neurological disorders [12],[13]. MAO exists in two isoforms, namely MAO A and MAO B. Serotonin (5-hydroxytryptamine) is preferentially oxidatively deaminated by MAO A while phenylethylamine and benzylamine are preferential substrates of MAO B for oxidative deamination [14]. Substrates common to both MAO isoforms include tryptamine, tyramine, and dopamine.

The three-dimensional structure MAO B has been described. It consists of 520 amino acids [15]. It has an FAD moiety covalently linked to a cysteine residue, Cys-397, through an 8α-(cysteinyl)-riboflavin linkage [16],[17],[18],[19]. In the structure of the enzyme, FAD-binding site, active site, and the regions that are involved in binding and specificities of the substrate and inhibitors have been described [20],[21],[22],[23]. Amino-acid residues like Lys-296, Trp-388, Tyr-398, and Tyr-435 appear to play important roles in the catalytic activity of MAO and may be involved in binding FAD as well via noncovalent linkages. An aromatic sandwich formed by Tyr-398 and Tyr-435 seems to stabilize the bound substrate. Other key amino-acid residues in the active site of the enzyme include Trp-119, Leu-164, Phe-168, Leu-171, Cys-172, Tyr-188, Ile-199, Gln-206, and Tyr-326. These residues may be involved in various interactions with the substrate/ligands including hydrophobic, hydrogen-bonding, and pi–pi interactions.

Molecular docking is a commonly used computational method that is used to study ligand–protein interactions. Several software programs are available in the public domain to perform small-molecule–protein docking and to analyze docking interactions. In this study, molecular docking program AutoDock 4.2 was used to dock ligands with MAO in an attempt to study the interactions. This is a first study of its kind involving active principles of khat, CATHI, and CATHO.

  Materials and methods Top

Molecular docking was performed by using the molecular docking software AutoDock, Version 4.2 [24]. For preparation of the protein and the ligands for docking, AutoDock Tools [24],[25], which is a graphic user interface for AutoDock 4.2, was used. The source of both the software was The Scripps Research Institutes (San Diego, California, USA).

Preparation of protein and ligand

PDB file ID 2V5Z was downloaded from the Protein Data Bank for the three-dimensional structure of subunit A of monoamine oxidase B (MAO B.A). Structures of ligands, safinamide (SAF), AMPH, CATHI, and CATHO were obtained as sdf files from the PubChem database and converted to pdb formats using Open Babel software. In the preparation of the protein and ligands for docking nonpolar hydrogens were merged and Gesteiger partial charges were assigned to all atoms. In applying torsions in ligands all rotatable bonds were rotated. Ligands were made flexible and protein was kept rigid. PDBQT files (file format that contains partial charges and torsion records along with atom coordinates) were written for protein and each ligand and were used as input files for docking experiments in the next step.


Standard docking procedures for a rigid protein and a flexible ligand were used as per the user guide for AutoDock 4.2. Briefly, using AutoGrid (a component of the software), a grid of 60×60×60 points in x, y, and z directions was built with grid points spaced at 0.375 Å. Electrostatic maps were calculated by using a distance-dependent function of the dielectric constant. All other parameters were set as default as per the user guide. Docking simulations were performed by using the Lamarckian genetic algorithm (as per the user guide). The implementation of Lamarckian genetic algorithm included creation of an initial population of 150 individuals. Random torsions were applied to each of 150 individuals. In each docking run, a maximum of 2 500 000 energy evaluations was performed. For each of four ligands, at least 20 such runs were performed. The best binding modes for each ligand obtained from docking were analyzed by using LigPlot+, AutoDock Tools, and RasMolR (Roger Sayle) [26] programs.

  Results and discussion Top

All binding parameters of SAF, AMPH, CATHI, and CATHO obtained after docking with MAO B.A are listed in [Table 1].
Table 1 Interaction energies and inhibitor constants (Ki) for the binding of safinamide, amphetamine, cathine, and cathinone with monoamine oxidase B A

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Estimated total free energy of binding of four inhibitors was −9.61, −6.25, −6.05, and −6.24 kcal/mol, respectively. The estimated Ki values were 9.08×10−8 M, 2.61×10−5 M, 3.69×10−5 M, and 2.69×10−5 M, respectively. The total free energy of binding (and hence the Ki) estimated for SAF is substantially lower than that of the other three ligands which suggests a strong affinity for this inhibitor. While binding affinities of AMPH, CATHI, and CATHO are almost equal to each other, these affinities are appreciably lower when compared with that of SAF. Inhibitor constants in the micromolar range for AMPH, CATHI, and CATHO indicate a weak inhibition of the enzyme while SAF is a known strong inhibitor of MAO B with an inhibitor constant in the nanomolar range.A structural rendering of the docked ligand–protein complex, depicting all four docked inhibitors and secondary structural features of MAO B.A, is shown in [Figure 1].
Figure 1 Three-dimensional structure of MAO B.A showing secondary structural details along with ligands bound in the active site. Alpha chains are in red, beta strands are in yellow, and ligands are shown overlapped in the active site. MAO B, monoamine oxidase B.

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Binding modes of AMPH, CATHI, and CATHO within the active site have been shown to be overlapped on each other in [Figure 2]. An analysis of docked complexes of all inhibitors was done using the LigPlot+ software. All LigPlots are shown in [Figure 3],[Figure 4],[Figure 5]. [Table 2] lists all significant interactions of ligands with active site residues of protein.
Figure 2 Conformations of AMPH, CATHI, and CATHO within the active site of MAO B.A shown overlapped. Red, AMPH; green, CATHI; Cyan, CATHO. AMPH, amphetamine; CATHI, cathine; CATHO, cathinone; MAO B, monoamine oxidase B.

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Figure 3 LigPlot of binding interaction of SAF in the active site of MAO B.A. Atoms are shown in CPK colors. H-bonds are shown as green dotted lines. Hydrophobic interactions are shown as red dotted lines and rays from residues toward ligand atoms. MAO B, monoamine oxidase B; SAF, safinamide.

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Figure 4 LigPlot of binding interaction of AMPH in the active site of MAO B.A. Atoms are shown in CPK colors. H-bonds are shown as green, dotted lines. Hydrophobic interactions are shown as red dotted lines and rays from residues toward ligand atoms. AMPH, amphetamine; MAO B, monoamine oxidase B.

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Figure 5 LigPlot of binding interaction of CATHI and CATHO in the active site of MAO B.A. Atoms are shown in CPK colors. H-bonds are shown as green, dotted lines. Hydrophobic interactions are shown as red, dotted lines and rays from residues toward ligand atoms. CATHI, cathine; CATHO, cathinone; MAO B, monoamine oxidase B.

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Table 2 Interactions of different active site residues of monoamine oxidase B A with ligand groups of safinamide, amphetamine, cathine, and cathinone

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Binding of SAF to MAO B has been described in detail earlier [27]. Binding mode of SAF with MAO B.A obtained in our docking experiments correspond well with those described. Binding of SAF with MAO B.A reveals several significant interactions of ligand within the active site of the enzyme. Extensive hydrophobic contacts are seen between ligand groups and active site residues on either side in active sites ([Figure 3]). The fluorobenzyl ring is in contact with residues Trp-119, Leu-164, Phe-168, Ile-199, and Tyr-326. The other benzyl ring that carries the terminal propanamide side chain makes hydrophobic contacts with residues Leu-171, Cys-172, Ile-199, and Gln-206. The terminal propanamide chain is involved in contacts with Cys-172, Tyr-398, and Tyr-435. A notable feature of the interaction of SAF with MAO B.A is the existence of a hydrogen bond between the phenyl O of Tyr-188 and the amido O of the propanamide side chain.

The binding modes of AMPH, CATHI, and CATHO are very similar to each other ([Figure 2]) and suggest that these inhibitors bind in the same site (active site) as SAF. Several interactions of these inhibitors with MAO B.A are comparable to the interactions of SAF ([Figure 3],[Figure 4],[Figure 5] and [Table 2]). The weaker binding of AMPH, CATHI, and CATHO compared with that of SAF is explained by the difference in their structure. While the fluorobenzyl ring in SAF allows greater extent of hydrophobic contacts, this additional ring is absent in three inhibitors. Hence, a weaker affinity and greater Ki. For three inhibitors, the benzyl ring makes contacts with residues Leu-171, Ile-198, Ile-199, Gln-206, and Tyr-326 while the propylamine side chain exhibits hydrophobic interactions with residues Leu-171, Tyr-398, and Tyr-435. A hydrogen bond seems to form between the backbone CO of Cys-172 and N of propylamine side chain. A notable difference in structures of CATHI and CATHO from that of AMPH is the presence of an additional alcohol O (in CATHI) and a ketone O (in CATHO). This oxygen does not appear to interact significantly. Owing to the structural similarity, all these three inhibitors show similar binding modes and affinities. The inhibition of MAO B caused by these inhibitors is weak on account of lesser affinity of binding. Yet, this inhibition of MAO B might be responsible for the psychoactivity of CATHI and CATHO (and hence khat). It may be speculated that since MAO B is involved in the metabolism of neurotransmitter amines, its inhibition might result in sustained levels of these neurotransmitters.

  Conclusion Top

CATHI and CATHO appear to bind to MAO B enzyme in silico. Their binding mode is similar to that of AMPH which has been shown to inhibit MAO B. The inhibitor constants obtained from the docking studies are in the micromolar range and very similar for AMPH, CATHI, and CATHO. Although, the three inhibitors show weak binding with MAO when compared with that of a strong inhibitor, SAF (nanomolar inhibitor constant), such inhibition may be significant. The psychoactivity associated with khat may be a result of sustained levels of neurotransmitter amines that may result due to the inhibition of MAO B by CATHI and CATHO (as MAO is responsible for the metabolism of these neurotransmitters). The binding of CATHI and CATHO with MAO B was studied using molecular docking, but it may be interesting to study such inhibition in vitro. It will also be of interest to see if CATHI and CATHO inhibit other enzymes in silico and/or in vitro.

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Conflicts of interest

There are no conflcits of interest.

  References Top

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  [Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5]

  [Table 1], [Table 2]


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