2548 Journal of Medicinal Chemistry, 2009, Vol. 52, No 8 Paz et al was made because, as discussed, the electron density there is (7) Harvey, A L. The phar quite poor, most likely due to local disorder and the rmsd Pharmacol. Ther. 1995. 68 between the 5h from the crystal structure and the models was (8)Zhang R. w: Tang, X C; Han, Y.Y.; Sang, G. W i Zhang, Y D. Ma, Y.X.: Zhang, C L; Yang, R. M. Drug calculated only for the CAS MEP, linker region, and the nitrogen A in the treatment of senile memory disorders. Acta Pharmacol. Si of the CAs MEP Most of the different gold models obtained 1991.12.250-252 for this complex, using different AChE molecules(apo and holo (9)Alvarez, A: Bronfman, F: Perez, C.A. Vicente, M: Garrido, J enzymes, Tc and mouse), share an overall similarity but many, plaque component. brillogenesis of amyloid-beta-peptides. Neurosci. Left in detail, fail to capture the exact nuances of the 5h-TCAChE 1995,201,49-52. Interactions (10)Alvarez, A Opazo, C. Alarcon, R; Garrido, J; Inestrosa, N. C Acetylcholinesterase promotes the aggregation of amyloid-beta-peptio phenol oriented down the gorge toward the catalytic③3mmm吗用M业B Since an earlier modeling study of a MEP/TcAChE compl N.C∴ Alvarez,A; Perez foreno. R. D. 1.: Linker. C: Casanueva. O. I: Soto, C: Garrido, J. Acet the spacers attached via the azepane rings and not through the imer's fibrils: possible role of the peripheral site of the enzym oxygen atoms of the phenol groups. 9 The crystal structure of 1996,16,881-891 the 5h/TcAChE complex described here displays a somewhat (12)Bartolini, M.; Bertucci, C; Cavrini, V: Andrisano, V. beta-Amyloid similar orientation of mep within the cas but also reveals high aggregation induced by human acetylcholinesterase: inhibition studies. Biochem. Pharmacol. 2003. 65. 407- flexibility of the MEP moiety at the PAs. It might, therefore, (13) Reyes, A E. Perez, D. R. Alvarez, A: Garrido, J: Gentry, M.K. be worth considering use of another atom on the azepane ring Doctor, B. P. Inestrosa. N. C. A me for linkage to the spacer or even synthesizing a molecule in acetylcholinesterase inhibits the formation of amyloid fibrils induce by the enzyme. Biochem. Biophys. Res. Commun. 1997, 232, 652- and the other through its azepane ring. These derivatives might (14)Sussman, J. Li; Harel, M. Frolow, F; Oefner, C; Goldman, A. have different affinities and specificities for AChE and possibl L; Silman, I. Atomic structure of acetylcholinesterase from To ind more specifically to the PAS than 5h. This could provide alifomica: a prototypic acetylcholine-binding protein Scien 253,872-879 information as to whether flexible binding of moieties at the (15)Silma PAS might affect the inhibitory effect on AB deposition. Acknowledgment. This work was supported in part by the T grants by the Divadol Foundation, the Israel Science Foundation nd low cost bis-tetrahydroaminacrine inhibitors of acetylcholinesterase. Steps toward novel drugs for treating Alzheimers the Nalvyco Foundation, the Neuman Foundation, the Bruce disease.J.Biol.chem.1996.271,23646-23649 rosen foundation the jean and Jula Goldwurm Memorial (17) Du, D M; Carlier, P R Development Foundation, a research grant from Erwin Pearl, the Benoziyo drugs for Alzhe acetylcholinesterase Pham.Des.2004,lO,3141-3156 Center for Neuroscience, the European Commission Sixth (18)Haviv, H :W D. M.: Silman. I: Sussman. J. L. Bivalent ligands Framework Research and Technological Development Pro- derived from Huperzine A tylcholinesterase inhibitors. Curr. ramme"SPINE2-COMPLEXES project under contract num- Top.Med.Chem.2007,7,375-387 ber LSHG-CT-2006-031220, Teach-SG" project under contract (19)Xie, Q; Wang, H; Xia, Z; Lu, M: Tang. Y. Sheng. W: Li. W: Zhou. W: Zhu, number ISSG-CT-2007-037198 (I.S. and J. LS), and by grants ptazinols as novel nanomolar ch from the national natural science foundation of pr china with high inhibitory potency on amyloid-beta (nos. 30472088 and 30772553)and the Chinese lll Project to ECUST. J L.S. is the Morton and Gladys Pickman (20) Jones, G: Willett, P: Glen, R. C: Leach, A. R: Taylor, R ation of a genetic algorithm for flexible docking Professor of Structural Biology. We are grateful to Esther Roth J.Mol.Biol.1997.267.727-7 and Lilly Toker for the samples of purified TcAChE. (21) an J. L; HareL, M. Frolow. F: Varon, L; Toker. L: Futerman man, I. Purification and crystallization of a dimeric form of Supporting Information Available: Structures acetylcholinesterase from Torpedo californica subsequent to solubi (ZIP): wbc_b_ee_447-pdb, wbc_b_ea_220 pdb, 2wbc Biol.1988,203,821-82 2wbc_leas_ee_531. pdb, wbc_leasea_52l pdb, 2wbc (22)McPherson, A Preparation and Analysis of Protein Crystals; John ThismaterialisavailablefreeofchargeviatheInternetathttp:// Wiley Sons: New York, 1982 pubs. acs. org 23)Otwinowski, Z. Minor, w. Processing of X-ray diffraction data collected in oscillation mode Methods Enzymol. 1997. 276. 307-326 (24)CCP4. The CCP4 suite: ns for protein crystallography. Acta References llgr.1994,50,760-7 (25) Erera io. A icons Kilo. Set at ot of cergstilogin ngis. Mol.Biol.1975.43.103-218 (2)Bartus, R. T: Dean, R. L, Ill; Beer, B: Lippa, A. S. The cholinergic hypothesis of geriatric memory dysfunction. Science 1982, 217, 408 (26)Walters, P. Stahl, M., Babel, version 1.1: Department of Chemistry, niversity of Arizona: Tucson, AZ, 1994 (27)Emsley, P: Cowtan, K. Coot: Model-Building Tools for Moled (3)Dunnett, S. B. Fibiger, H. C. Role of forebrain cholinergic systen Graphics. Acta Crystallogr., Sect. D: Biol. Crystallogr. 2004, 60, 2126- relevance to the cog (28)Davis, I. W: Leaver-Fay, A: Chen, V.B.; Block, J N Kapral, G.J. (4)Greenblatt, H. M: Dvir, H: Silman, L; Sussman, J. L. Acetylcho- inesterase: a multifaceted target for structu hardson, J.S.: Richardson, D. C. MolProbity: all-atom contacts anticholinesterase agents for the treatment of Alzheimers disease. and structure validation for proteins and nucleic acids. Nucleic Acids Mo. Neurosci.2003.20,369-383. Res.2007.35.W375-W383 (5)Kawakami, Y: Inoue, A: Kawai, T; Wakita, M: Sugimoto, H. (29) Adams, P D. Grosse-Kunstleve, Rw T.R. Hopfinger, A J. The rationale for E2020 as a potent acetylcholinest McCoy, A. J: Moriarty, N. w; Read, R. hettini. J C: Saut erase inhibitor. Bioorg. Med. Chem. 1996. 4. 1429-1446 PHENIX lg new software for (6)Enz, A: Boddeke, H; Gray, J; Spiegel, R linicopharmacologic properties of SDZ ENA 713, Sect. D: Biol. Crystallogr. 2002, 58, 1948-1954 cetylcholinesterase inhibitor. Ann. N.Y. Acad. Sc 0.272 (0)Chen, Y. Studies on the synthesis, resolution and optical isomers of 275 meptazinol. Dissertation. Fudan Univ: Shanghai, 2004: Pp 19-2was made because, as discussed, the electron density there is quite poor, most likely due to local disorder and the rmsd between the 5h from the crystal structure and the models was calculated only for the CAS MEP, linker region, and the nitrogen of the CAS MEP. Most of the different GOLD models obtained for this complex, using different AChE molecules (apo and holo enzymes, Tc and mouse), share an overall similarity but many, in detail, fail to capture the exact nuances of the 5h-TcAChE interactions. Since an earlier modeling study of a MEP/TcAChE complex placed the azepane ring in the middle of the gorge with the phenol oriented down the gorge toward the catalytic site,37 the bivalent MEP derivatives were designed and synthesized with the spacers attached via the azepane rings and not through the oxygen atoms of the phenol groups.19 The crystal structure of the 5h/TcAChE complex described here displays a somewhat similar orientation of MEP within the CAS but also reveals high flexibility of the MEP moiety at the PAS. It might, therefore, be worth considering use of another atom on the azepane ring for linkage to the spacer or even synthesizing a molecule in which one MEP would be connected through its phenol group and the other through its azepane ring. These derivatives might have different affinities and specificities for AChE and possibly bind more specifically to the PAS than 5h. This could provide information as to whether flexible binding of moieties at the PAS might affect the inhibitory effect on A
deposition. Acknowledgment. This work was supported in part by the grants by the Divadol Foundation, the Israel Science Foundation, the Nalvyco Foundation, the Neuman Foundation, the Bruce Rosen Foundation, the Jean and Jula Goldwurm Memorial Foundation, a research grant from Erwin Pearl, the Benoziyo Center for Neuroscience, the European Commission Sixth Framework Research and Technological Development Pro￾gramme “SPINE2-COMPLEXES” project under contract num￾ber LSHG-CT-2006-031220, “Teach-SG” project under contract number ISSG-CT-2007-037198 (I.S. and J.L.S.), and by grants from the National Natural Science Foundation of P.R. China (nos. 30472088 and 30772553) and the Chinese 111 Project grant to ECUST. J.L.S. is the Morton and Gladys Pickman Professor of Structural Biology. We are grateful to Esther Roth and Lilly Toker for the samples of purified TcAChE. 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