Determination by X-ray Crystallography of the Three-Dimensional Structure ofAcetylcholinesterase from Torpedo Electric Organ

摘要

The objective of this project is the determination of the three-dimensionalstructure of acetylcholinesterase (AChE) from electric organ tissue of Torpedo californica, with the intention of elucidating the detailed topography of its catalytic site. In the period covered by this report, work was carried on the dimeric form of AChE from the above source, purified by affinity chromatography subsequent to solubilization with phosphatidylinositol-specific phospholipase C, which contains two catalytic subunits (each of M(sub r) = 65,000) linked by a single disulfide bond. Crystals were obtained from this preparation by vapor diffusion, using ammonium sulfate as the precipitant. These crystals belong to space group p3(sub121) with unit cell dimensions a=b=110.9 A and c=136.9 A. Native x-ray data sets were collected on a Siemens/Xentronics Area Detector to 2.8 A resolution. A search for heavy-atom derivatives was conducted using six different heavy-atom compounds under a variety of soaking conditions. Soaking with 2 mili molar UO2 (NO3)2, in a phosphate-free mother liquor for 8 days, yielded an isomorphous heavy-atom derivative. Based on difference Patterson maps, it was possible to find two-major and two minor sites for this derivative.