Journal:Neuropharmacology:2

Computational Studies on Cholinesterases: Strengthening our Understanding of the Integration of Structure, Dynamics and Function

Joel L. Sussman and Israel Silman ^[1]

Molecular Tour
Computational approaches have proved valuable in elucidating structure/function relationships in the cholinesterases in the context of their unusual three-dimensional structure. In this review we survey several recent studies that have enhanced our understanding of how these enzymes function, and have utilized computational approaches both to modulate their activity and to improve the design of lead compounds for their inhibition.

Solution of the crystal structure of Torpedo californica acetylcholinesterase (TcAChE) in 1991 (Sussman et al., 1991)^[2], revealed a three-dimensional structure that was wholly unanticipated. Despite the fact that AChE is one of the most rapid enzymes known, operating at a speed approaching diffusion control (Bazelyansky et al., 1986;^[3] Rosenberry, 1975^[4]), its active site is deeply buried, at the bottom of a long and narrow gorge, whose cross-section, at its narrowest point, is significantly smaller than the cross-section of the quaternary group of acetylcholine (ACh). Subsequent solution of the crystal structures of mouse (m) (Bourne et al., 1995^[5]), Electrophorus electricus (Ee) (Bourne et al., 1999;^[6] Raves et al., 1998^[7]), human (h) (Cheung et al., 2012;^[8] Kryger et al., 2000^[9]), Drosophila melanogaster (Dm) (Harel et al., 2000^[10]), Bungarus fasciatus (Bf) (Bourne et al., 2015^[11]), and Anopheles gambiae (Ag) (Cheung et al., 2018;^[12] Han et al., 2018^[13]) AChEs, as well as that of human serum butyrylcholinesterase (hBChE) (Nicolet et al., 2003^[14]), revealed highly homologous structures. The crystal structures of the cholinesterases (ChEs) raised cogent questions with respect to the coupling of the structure of the enzymes, together with their dynamics, to their catalytic activity, and to the way in which inhibitors, some of which are large and rigid, bind to, and disassociate from them. These topics have been covered extensively in earlier reviews (Silman and Sussman, 2008;^[15] Xu et al., 2017^[16]). However, as computing power has continued to grow, databases have expanded, and increasingly sophisticated algorithms have been developed, problems become accessible that had previously seemed unapproachable (Fuxreiter, 2015^[17]). In the following, after briefly surveying the structure and dynamics of the ChEs, taking TcAChE as the prototypic case, we wish to briefly present and discuss some recent studies that cover various aspects of these topics.

, and expressed in E coli. It is displayed in ribbon form, colored from the N-terminus to the C-terminus in a spectrum going from blue to red. The 51 amino acids that were mutated on the basis of the prediction of the PROSS algorithm are shown as magenta spheres.

. Carbon atoms of H438 and E325 are shown as green sticks for the wild-type conformation. Carbon atoms of H438 and E322, as well as of the oxyanion hole residues - G116, G117 and A199 - are shown as cyan sticks for the mutant. In both cases, oxygens are coded in red, and nitrogens in blue. The OP moiety covalently attached to S198 in the mutant is displayed as balls, with the phosphorus in orange, the oxygens in red and the carbons in cyan. H-bonds are shown as white dashed lines.

Docking and MD simulation for interaction of BSF and PMSF with TcAChE and mAChE. In all four scenes two copies of the ligand are displayed. One shows the position of the ligand after docking alone (blue), and the other shows the position after docking followed by MD simulation (orange). It should be noted that the orientations of the amino-acid side-chains displayed are those seen prior to the MD simulations.

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. Three PEGs are shown in red, methylene blue in blue, and conserved aromatic residues lining the active-site gorge in green. A highly conserved H₂O molecule, shown as a yellow sphere, also affects the positioning of the ligand.

Active-site gorge in the and structures of native DmAChE. Residues of the catalytic triad (E367/H480/S238), of the oxyanion hole (G150/G151/Al239), and key residues of the peripheral site (W321), acyl-binding pocket (W271), and choline-binding pocket (W83), are represented as sticks, with carbons in white, nitrogens in blue, and oxygens in red. The alternative conformation of W83 is depicted with carbons in cyan. The acetyl (ACT) is represented as balls and sticks. H-bonds are depicted as black dashes, with distances in Å.

References

1. ↑ Sussman JL, Silman I. Computational studies on cholinesterases: Strengthening our understanding of the integration of structure, dynamics and function. Neuropharmacology. 2020 Aug 11;179:108265. doi: 10.1016/j.neuropharm.2020.108265. PMID:32795461 doi:http://dx.doi.org/10.1016/j.neuropharm.2020.108265
2. ↑ Sussman JL, Harel M, Frolow F, Oefner C, Goldman A, Toker L, Silman I. Atomic structure of acetylcholinesterase from Torpedo californica: a prototypic acetylcholine-binding protein. Science. 1991 Aug 23;253(5022):872-9. PMID:1678899
3. ↑ Bazelyansky M, Robey E, Kirsch JF. Fractional diffusion-limited component of reactions catalyzed by acetylcholinesterase. Biochemistry. 1986 Jan 14;25(1):125-30. doi: 10.1021/bi00349a019. PMID:3954986 doi:http://dx.doi.org/10.1021/bi00349a019
4. ↑ Rosenberry TL. Acetylcholinesterase. Adv Enzymol Relat Areas Mol Biol. 1975;43:103-218. doi:, 10.1002/9780470122884.ch3. PMID:891 doi:http://dx.doi.org/10.1002/9780470122884.ch3
5. ↑ Bourne Y, Taylor P, Marchot P. Acetylcholinesterase inhibition by fasciculin: crystal structure of the complex. Cell. 1995 Nov 3;83(3):503-12. PMID:8521480
6. ↑ Bourne Y, Grassi J, Bougis PE, Marchot P. Conformational flexibility of the acetylcholinesterase tetramer suggested by x-ray crystallography. J Biol Chem. 1999 Oct 22;274(43):30370-6. PMID:10521413
7. ↑ doi: https://dx.doi.org/10.1007/978-1-4899-1540-5_97
8. ↑ Cheung J, Rudolph MJ, Burshteyn F, Cassidy MS, Gary EN, Love J, Franklin MC, Height JJ. Structures of human acetylcholinesterase in complex with pharmacologically important ligands. J Med Chem. 2012 Oct 4. PMID:23035744 doi:http://dx.doi.org/10.1021/jm300871x
9. ↑ Kryger G, Harel M, Giles K, Toker L, Velan B, Lazar A, Kronman C, Barak D, Ariel N, Shafferman A, Silman I, Sussman JL. Structures of recombinant native and E202Q mutant human acetylcholinesterase complexed with the snake-venom toxin fasciculin-II. Acta Crystallogr D Biol Crystallogr. 2000 Nov;56(Pt 11):1385-94. PMID:11053835
10. ↑ Harel M, Kryger G, Rosenberry TL, Mallender WD, Lewis T, Fletcher RJ, Guss JM, Silman I, Sussman JL. Three-dimensional structures of Drosophila melanogaster acetylcholinesterase and of its complexes with two potent inhibitors. Protein Sci. 2000 Jun;9(6):1063-72. PMID:10892800
11. ↑ Bourne Y, Renault L, Marchot P. Crystal Structure of Snake Venom Acetylcholinesterase in Complex with Inhibitory Antibody Fragment Fab410 bound at the Peripheral Site: Evidence for Open and Closed States of a Backdoor Channel. J Biol Chem. 2014 Nov 19. pii: jbc.M114.603902. PMID:25411244 doi:http://dx.doi.org/10.1074/jbc.M114.603902
12. ↑ Cheung J, Mahmood A, Kalathur R, Liu L, Carlier PR. Structure of the G119S Mutant Acetylcholinesterase of the Malaria Vector Anopheles gambiae Reveals Basis of Insecticide Resistance. Structure. 2018 Jan 2;26(1):130-136.e2. doi: 10.1016/j.str.2017.11.021. Epub 2017, Dec 21. PMID:29276037 doi:http://dx.doi.org/10.1016/j.str.2017.11.021
13. ↑ Han Q, Wong DM, Robinson H, Ding H, Lam PC, Totrov MM, Carlier PR, Li J. Crystal structure of acetylcholinesterase catalytic subunits of the malaria vector Anopheles gambiae. Insect Sci. 2017 Mar 1. doi: 10.1111/1744-7917.12450. PMID:28247978 doi:http://dx.doi.org/10.1111/1744-7917.12450
14. ↑ Nicolet Y, Lockridge O, Masson P, Fontecilla-Camps JC, Nachon F. Crystal structure of human butyrylcholinesterase and of its complexes with substrate and products. J Biol Chem. 2003 Oct 17;278(42):41141-7. Epub 2003 Jul 17. PMID:12869558 doi:http://dx.doi.org/10.1074/jbc.M210241200
15. ↑ Silman I, Sussman JL. Acetylcholinesterase: how is structure related to function? Chem Biol Interact. 2008 Sep 25;175(1-3):3-10. Epub 2008 Jun 6. PMID:18586019 doi:10.1016/j.cbi.2008.05.035
16. ↑ Xu Y, Cheng S, Sussman JL, Silman I, Jiang H. Computational Studies on Acetylcholinesterases. Molecules. 2017 Aug 10;22(8). pii: molecules22081324. doi:, 10.3390/molecules22081324. PMID:28796192 doi:http://dx.doi.org/10.3390/molecules22081324
17. ↑ doi: https://dx.doi.org/10.1201/b17979