title: The impact of crystallization conditions on structure-based drug design: A case study on the methylene blue/acetylcholinesterase complex.
abstract: Structure-based drug design utilizes apoprotein or complex structures retrieved from the PDB. >57% of crystallographic PDB entries were obtained with polyethylene glycols (PEGs) as precipitant and/or as cryoprotectant, but <6% of these report presence of individual ethyleneglycol oligomers. We report a case in which ethyleneglycol oligomers' presence in a crystal structure markedly affected the bound ligand's position. Specifically, we compared the positions of methylene blue and decamethonium in acetylcholinesterase complexes obtained using isomorphous crystals precipitated with PEG200 or ammonium sulfate. The ligands' positions within the active-site gorge in complexes obtained using PEG200 are influenced by presence of ethyleneglycol oligomers in both cases bound to W84 at the gorge's bottom, preventing interaction of the ligand's proximal quaternary group with its indole. Consequently, both ligands are ∼3.0Å further up the gorge than in complexes obtained using crystals precipitated with ammonium sulfate, in which the quaternary groups make direct π-cation interactions with the indole. These findings have implications for structure-based drug design, since data for ligand-protein complexes with polyethylene glycol as precipitant may not reflect the ligand's position in its absence, and could result in selecting incorrect drug discovery leads. Docking methylene blue into the structure obtained with PEG200, but omitting the ethyleneglycols, yields results agreeing poorly with the crystal structure; excellent agreement is obtained if they are included. Many proteins display features in which precipitants might lodge. It will be important to investigate presence of precipitants in published crystal structures, and whether it has resulted in misinterpreting electron density maps, adversely affecting drug design.
authors.affiliation: Israel Structural Proteomics Center, Weizmann Institute of Science, Rehovot, 76100, Israel.|Department of Biological Chemistry, Weizmann Institute of Science, Rehovot, 76100, Israel.
authors.forename: Orly
authors.initials: O
authors.lastname: Dym
authors.affiliation: CAS Key Laboratory of Receptor Research, Drug Discovery and Design Center, Shanghai Institute of Materia Medica, Chinese Academy of Sciences (CAS), Shanghai (22), China.
authors.forename: Wanling
authors.initials: W
authors.lastname: Song
authors.affiliation: Department of Structural Biology, Weizmann Institute of Science, Rehovot, 76100, Israel.
authors.forename: Clifford
authors.initials: C
authors.lastname: Felder
authors.affiliation: Department of Neurobiology, Weizmann Institute of Science, Rehovot, 76100, Israel.
authors.forename: Esther
authors.initials: E
authors.lastname: Roth
authors.affiliation: Department of Biochemistry and Molecular Biology, Universidad de Salamanca, Salamanca, 37007, Spain.
authors.forename: Valery
authors.initials: V
authors.lastname: Shnyrov
authors.affiliation: Department of Structural Biology, Weizmann Institute of Science, Rehovot, 76100, Israel.
authors.forename: Yacov
authors.initials: Y
authors.lastname: Ashani
authors.affiliation: CAS Key Laboratory of Receptor Research, Drug Discovery and Design Center, Shanghai Institute of Materia Medica, Chinese Academy of Sciences (CAS), Shanghai (22), China.
authors.forename: Yechun
authors.initials: Y
authors.lastname: Xu
authors.affiliation: Department of Biochemistry, Netherlands Cancer Institute, Amsterdam, CX, 1066, the Netherlands.
authors.forename: Robbie P
authors.initials: RP
authors.lastname: Joosten
authors.affiliation: Department of Chemical Research Support, Weizmann Institute of Science, Rehovot, 76100, Israel.
authors.forename: Lev
authors.initials: L
authors.lastname: Weiner
authors.affiliation: Israel Structural Proteomics Center, Weizmann Institute of Science, Rehovot, 76100, Israel.|Department of Structural Biology, Weizmann Institute of Science, Rehovot, 76100, Israel.
authors.forename: Joel L
authors.initials: JL
authors.lastname: Sussman
authors.affiliation: Department of Neurobiology, Weizmann Institute of Science, Rehovot, 76100, Israel.
authors.forename: Israel
authors.initials: I
authors.lastname: Silman
mesh_terms: D000110:Acetylcholinesterase / Q000737:chemistry*; D000818:Animals; D015195:Drug Design; D029941:Fish Proteins / Q000737:chemistry*; D008751:Methylene Blue / Q000737:chemistry*; D062105:Molecular Docking Simulation; D014101:Torpedo
chemical_list: D029941:Fish Proteins; D000110:Acetylcholinesterase; D008751:Methylene Blue
keywords: acetylcholinesterase; decamethonium; methylene blue; polyethylene glycol; structure-based drug discovery