Taspine: bioactivity-guided isolation and molecular ligand-target insight of a potent acetylcholinesterase inhibitor from Magnolia x soulangiana

Abstract

A bioactivity-guided approach was taken to identify the acetylcholinesterase (AChE, EC 3.1.1.7) inhibitory agent in a Magnolia x soulangiana extract using a microplate enzyme assay with Ellman's reagent. This permitted the isolation of the alkaloids taspine (1) and (-)-asimilobine (2), which were detected for the first time in this species. Compound 1 showed a significantly higher effect on AChE than the positive control galanthamine and selectively inhibited the enzyme in a long-lasting and concentration-dependent fashion with an IC(50) value of 0.33 +/- 0.07 muM. Extensive molecular docking studies were performed with human and Torpedo californica-AChE employing Gold software to rationalize the binding interaction. The results suggested ligand 1 to bind in an alternative binding orientation when compared to galanthamine. While this is located in close vicinity to the catalytic amino acid triad, the 1-AChE complex was found to be stabilized by (i) sandwich-like pi-stacking interactions between the planar aromatic ligand (1) and the Trp84 and Phe330 of the enzyme, (ii) an esteratic site anchoring with the amino side chain, and (iii) a hydrogen-bonding network.

Figure 1

Microplate assay: inhibitory effects of taspine (1) and galanthamine on AChE. Statistical analysis: ***p < 0.001, **p < 0.01, *p < 0.05. Student’s t-test of absorption data after 30 min in comparison with medium control, n = 4.

Figure 2

Overlay of ligand-binding domains of hAChE (PDB: 1B41, stick style, highlighted) and TcAChE (PDBs: 1W6R, line style; 1ACJ, stick style) visualized with LigandScout. Galanthamine (from 1W6R) is shown in ball-and-stick style forming hydrogen-bonding interactions with two of the catalytic amino acids.

Figure 3

Two perspectives of tacrine bound to TcAChE (A and B) and two perspectives of taspine (1) docked into the bottom cavity of TcAChE revealing a binding mode comparable to tacrine (C and D) visualized with LigandScout. Color code: violet, π–π interactions; blue, positively ionized feature; green, hydrogen bond donor; red, hydrogen bond acceptor.