AChE inhibitors and substrates

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Revision as of 11:35, 1 July 2010

Dear readers, this page presents only a small part of the great world of the acetylcholinesterase inhibitors. So, please see also our pages AChE inhibitors and substrates (Part II), AChE inhibitors and substrates (Part III), AChE bivalent inhibitors and AChE bivalent inhibitors (Part II).

AChE substrate

Solution of the three-dimensional (3D) structure of Torpedo californica acetylcholinesterase (TcAChE) in 1991 (Sussman et al. & Silman (1991)) opened up new horizons in research on an enzyme that had already been the subject of intensive investigation. The unanticipated structure of this extremely rapid enzyme, in which the active site was found to be buried at the bottom of a , lined by (colored dark magenta), led to a revision of the views then held concerning substrate traffic, recognition and hydrolysis (Botti et al. Sussman & Silman (1999)). This led to a series of theoretical and experimental studies, which took advantage of recent advances in theoretical techniques for treatment of proteins, such as molecular dynamics and electrostatics and to site-directed mutagenesis, utilizing suitable expression systems. Acetylcholinesterase hydrolysizes the neurotransmitter acetylcholine , producing group. directly binds (via its nucleophilic Oγ atom) within the catalytic triad (ACh/TcAChE structure 2ace). The residues are also important in the ligand recognition .

AChE monovalent inhibitors (Part I)

Organophosphorus acid anhydride nerve agents

Organophosphorus (OP) acid anhydride nerve agents are potent inhibitors which rapidly phosphonylate AChE and then may undergo an internal dealkylation reaction (called "aging") to produce an OP-enzyme conjugate that cannot be reactivated.

Reaction between Ser200Oγ and Soman, assuming an in-line attack by the Oγ, followed by spontaneous dealkylation of the O-pinacolyl group.

As was mentioned above, AChE hydrolysizes the neurotransmitter , producing group. directly binds catalytic (via its nucleophilic Oγ atom). , O-(1,2,2-trimethylpropyl) methylphosphonofluoridate (fluorine atom is colored violet and phosphorus atom is colored darkmagenta), is one of the most toxic OPs. Soman inhibits AChE by to catalytic Ser200, . This process implicates nucleophilic attack of the Ser200 nucleophilic Oγ atom on the phosphorus atom of soman, with concomitant departure of its fluoride atom. After that AChE catalyzes the of the soman or other OP. This causes irreversible inhibition of AChE, "aged" soman/AChE conjugate can not be reactivated. However, before “aging”, at the step of , AChE can be by nucleophiles, such as pralidoxime (2-PAM), resulting in of the phosphonyl adduct from Ser200 Oγ. At the (2wfz) the catalytic His440 forms hydrogen bonds with Ser200 Oγ and Glu327 Oε1 via its Nε2 and Nδ1 nitrogens, respectively. The O2 atom of soman is within hydrogen bonding distance of His440 Nε2. Soman O1 mimicks carbonyl oxygen of ACh. A water molecule 1001 interacting with soman O2 is represented as a red ball. The active site residues of the nonaged soman/TcAChE are colored yellow. The O2 atom of the (2wg0) forms a salt bridge with His440 Nε2. The active site residues of the aged soman/TcAChE are colored pink. of the structures of the nonaged (2wfz) and aged (2wg0) conjugates reveals a small, but important, change within the active site - the imidazole ring of His440 is tilted back to a native-like conformation after dealkylation. The water molecule 1001, which interacts with soman O2 in the nonaged crystal structure, is not within hydrogen bonding distance of O2 in the aged crystal structure. 2-PAM binds poorly to the nonaged phosphonylated enzyme (its electron density was not found) and binds in an after soman aging to TcAChE (2wg1).

To understand the basis for irreversible inhibition, the structure of the aged conjugate obtained by reaction of TcAChE with soman was solved by X-ray crystallography to 2.2Å resolution (1som). The highest positive difference density peak corresponded to the OP phosphorus and was located within covalent bonding distance of the active-site serine (S200). The are within hydrogen-bonding distance of four potential donors from catalytic subsites of the enzyme, suggesting that electrostatic forces significantly stabilize the aged enzyme. The methyl group of soman occupies the , bounded by Trp233, Phe288, and Phe290.

Sarin, O-isopropylmethylphosponofluoridate, is an other toxic OP compound. It is also inhibits AChE by covalent binding to the catalytic Ser200. The active sites of aged (1cfj) and aged soman-TcAChE (1som and 2wg0) are almost identical and provided structural models for the negatively charged, tetrahedral intermediate that occurs during deacylation with the ACh. There are four hydrogen bond donors (red dotted lines) to the anionic phosphonyl oxygen atoms: the backbone amide nitrogen atoms of Ala201, Gly118, and Gly119, as well as His440 Nε2. The sarin methyl carbon (colored cyan) is within non-bonded contact distances (black dotted lines) of Phe288 and Phe290 in the acyl binding pocket.

DFP, diisopropylphosphorofluoridate, is an other toxic OP nerve agent. It is also inhibits AChE by covalent binding to the catalytic Ser200. As in the case with soman (1som) and sarin (1cfj), there are four hydrogen bond donors (dotted lines) to the anionic phosphonyl oxygen atoms: the backbone amide nitrogen atoms of Ala201, Gly118, and Gly119, as well as His440 Nε2 at the of aged DFP-TcAChE (2dfp). Phosphorylation with DFP caused an unexpected distortion in the main chain of a loop that includes residues F288 and F290 of the TcAChE acyl binding pocket. F288 and F290 move significantly in the DFP-TcAChE structure (lime), in comparison to their positions in the native enzyme (2ace). This is the first major conformational change reported in the active site of any AChE−ligand complex, and it offers a structural explanation for the substrate selectivity of AChE.

For information about additional AChE monovalent inhibitors please see AChE inhibitors and substrates (Part II).

AChE bivalent inhibitors

Please see pages AChE bivalent inhibitors and AChE bivalent inhibitors (Part II)

Selected 3D Structures of AChE

• 2ace This is the original solved structure for Torpedo Californica
• 1ea5 This is one of the highest quality representative X-ray structures in the PDB.
• 1eve The E2020 (Aricept) complex.
• 1ax9 Endrophonium complex.
• 1vot Complex with Huperzine, a Chinese folk medicine.
• 1fss Complex with snake venum toxin Fasciculin-II.
• 1acj Complex with tacrine.
• 1e66 Complex with huprine X.
• 1dx6 Complex with galanthamine.
• 1qti Complex with galanthamine.
• 1w6r Complex with galanthamine iminium derivative.
• 2ack Complex with edrophonium.
• 1vzj Structure of the tetramerization domain of acetylcholinesterase.
• 1gqr Complex with rivastigmine.
• 1gqs Complex with NAP alone.
• 1vxr Complex with VX.
• 2vja Complex with OTMA.
• 1som Complex with soman.
• 2wfz Complex with nonaged soman.
• 2wg0 Complex with aged soman.
• 2wg1 Complex with aged soman and 2-PAM.
• 1cfj Complex with aged sarin.
• 2dfp Complex with aged DFP.

More structures can be obtained by searching for AChE

References

• Raves ML, Harel M, Pang YP, Silman I, Kozikowski AP, Sussman JL. Structure of acetylcholinesterase complexed with the nootropic alkaloid, (-)-huperzine A. Nat Struct Biol. 1997 Jan;4(1):57-63. PMID:8989325
• Harel M, Schalk I, Ehret-Sabatier L, Bouet F, Goeldner M, Hirth C, Axelsen PH, Silman I, Sussman JL. Quaternary ligand binding to aromatic residues in the active-site gorge of acetylcholinesterase. Proc Natl Acad Sci U S A. 1993 Oct 1;90(19):9031-5. PMID:8415649
• Greenblatt HM, Guillou C, Guenard D, Argaman A, Botti S, Badet B, Thal C, Silman I, Sussman JL. The complex of a bivalent derivative of galanthamine with torpedo acetylcholinesterase displays drastic deformation of the active-site gorge: implications for structure-based drug design. J Am Chem Soc. 2004 Dec 1;126(47):15405-11. PMID:15563167 doi:http://dx.doi.org/10.1021/ja0466154
• Ravelli RB, Raves ML, Ren Z, Bourgeois D, Roth M, Kroon J, Silman I, Sussman JL. Static Laue diffraction studies on acetylcholinesterase. Acta Crystallogr D Biol Crystallogr. 1998 Nov 1;54(Pt 6 Pt 2):1359-66. PMID:10089512
• Haviv H, Wong DM, Greenblatt HM, Carlier PR, Pang YP, Silman I, Sussman JL. Crystal packing mediates enantioselective ligand recognition at the peripheral site of acetylcholinesterase. J Am Chem Soc. 2005 Aug 10;127(31):11029-36. PMID:16076210 doi:http://dx.doi.org/10.1021/ja051765f
• Carlier PR, Du DM, Han Y, Liu J, Pang YP. Potent, easily synthesized huperzine A-tacrine hybrid acetylcholinesterase inhibitors. Bioorg Med Chem Lett. 1999 Aug 16;9(16):2335-8. PMID:10476864
• Wong DM, Greenblatt HM, Dvir H, Carlier PR, Han YF, Pang YP, Silman I, Sussman JL. Acetylcholinesterase complexed with bivalent ligands related to huperzine a: experimental evidence for species-dependent protein-ligand complementarity. J Am Chem Soc. 2003 Jan 15;125(2):363-73. PMID:12517147 doi:http://dx.doi.org/10.1021/ja021111w
• 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
• Greenblatt HM, Kryger G, Lewis T, Silman I, Sussman JL. Structure of acetylcholinesterase complexed with (-)-galanthamine at 2.3 A resolution. FEBS Lett. 1999 Dec 17;463(3):321-6. PMID:10606746
• Bar-On P, Millard CB, Harel M, Dvir H, Enz A, Sussman JL, Silman I. Kinetic and structural studies on the interaction of cholinesterases with the anti-Alzheimer drug rivastigmine. Biochemistry. 2002 Mar 19;41(11):3555-64. PMID:11888271
• Colletier JP, Bourgeois D, Sanson B, Fournier D, Sussman JL, Silman I, Weik M. Shoot-and-Trap: use of specific x-ray damage to study structural protein dynamics by temperature-controlled cryo-crystallography. Proc Natl Acad Sci U S A. 2008 Aug 19;105(33):11742-7. Epub 2008 Aug 13. PMID:18701720
• Dvir H, Wong DM, Harel M, Barril X, Orozco M, Luque FJ, Munoz-Torrero D, Camps P, Rosenberry TL, Silman I, Sussman JL. 3D structure of Torpedo californica acetylcholinesterase complexed with huprine X at 2.1 A resolution: kinetic and molecular dynamic correlates. Biochemistry. 2002 Mar 5;41(9):2970-81. PMID:11863435
• Millard CB, Kryger G, Ordentlich A, Greenblatt HM, Harel M, Raves ML, Segall Y, Barak D, Shafferman A, Silman I, Sussman JL. Crystal structures of aged phosphonylated acetylcholinesterase: nerve agent reaction products at the atomic level. Biochemistry. 1999 Jun 1;38(22):7032-9. PMID:10353814 doi:http://dx.doi.org/10.1021/bi982678l
• Sanson B, Nachon F, Colletier JP, Froment MT, Toker L, Greenblatt HM, Sussman JL, Ashani Y, Masson P, Silman I, Weik M. Crystallographic Snapshots of Nonaged and Aged Conjugates of Soman with Acetylcholinesterase, and of a Ternary Complex of the Aged Conjugate with Pralidoxime (dagger). J Med Chem. 2009 Jul 30. PMID:19642642 doi:10.1021/jm900433t