The Cholinesterases—Structure, Mechanism, Function and Drug Design: The 25th Anniversary of the Solution of the Crystal Structure of Acetylcholinesterase by Joel L. Sussman and Israel Silman

Dear Colleagues,

Solution of the crystal structure of acetylcholinesterase (AChE) using an atomic resolution, in 1991, by Professors Joel Sussman and Israel Silman, revolutionized our understanding of structure/function relationships in this enzyme. Not only the structure itself, but also the concepts that they developed, stimulated major research efforts, both fundamental and applied, that come not only from the laboratories of these two scientists, but also from many other groups worldwide. The importance of their contribution is testified to by the fact that their original publication in Science in 1991 has already been cited more than 2700 times.

Prior to initiation of their collaboration, in 1985, both had already made seminal discoveries. Prof Israel Silman returned to the Weizmann Institute in Israel, in 1968, from a postdoctoral fellowship in the laboratory of Prof David Nachmansohn (Columbia University, New York), where he had initiated his studies on structure-function relationships in proteins of the cholinergic system, a theme that remains his principal focus. Back in Israel, Prof Silman devoted his main efforts to purification and characterization of AChE from electric organ tissue of the electric eel, Electrophorus electricus (Ee), and the electric ray, Torpedo californica (Tc). His notable achievements included, initially, characterization of the collagen-tailed asymmetric forms of the enzyme. Subsequently, he characterized the glycophosphatidylinositol (GPI)-anchored form of AChE, the first member of the large class of GPI-anchored proteins to be identified. This proved to be an essential first step in the eventual crystallization of TcAChE in 1988. Prof. Sussman is amongst the pioneers of macromolecular refinement, developing the CORELS program, and applying it to structural studies on yeast tRNA^phe. After moving to the Weizmann Institute from Duke University in North Carolina, in 1976, he continued his studies on nucleic acids, determining the structure of 'bulge'-containing DNA fragments, models for insertion mutations, and also worked on the structures of halophilic proteins isolated from bacteria that grow in the Dead Sea. He was a pioneer in developing methods of flash freezing protein and nucleic acid molecules to cryogenic temperatures in order to greatly prolong their lifetime under X-ray irradiation. He was the Director of the Protein Data Bank at Brookhaven National Laboratory from 1994–1999, where he helped to transform it into an interactive resource on the internet, to aid researchers in visualizing the structure/function relationships of biological macromolecules. The major focus of his work, since the late 1980s, has been on proteins of the nervous system in particular on AChE.

Profs. Silman and Sussman chose to initiate their collaborative effort by focusing on TcAChE, after unsuccessful attempts by others to solve the crystal structure of the 11S tetramer from EeAChE. The TcAChE dimer has a simpler quaternary structure than that of the EeAChE tetramer, and unlike it, the integrity of the TcAChE catalytic subunit is well preserved during a mild isolation and purification procedure. Solution of the 3D structure of TcAChE, and their continuing collaborative endeavors, have provided answers to numerous questions that had previously puzzled those working on AChE, and also established conceptual breakthroughs that gave both theoreticians and experimentalists a totally new perspective on the function of AChE itself, and on the cholinergic synapse. Their findings have not only had major ramifications in their particular area of research, but have also impacted greatly upon other fields. Sussman and Silman went on to solve the 3D structures of many complexes of AChE with reversible ligands—such as edrophonium, tacrine, donepezil (E2020), huperzine A and methylene blue—and of covalent conjugates with soman, sarin, VX, and rivastigmine. They elucidated the structural and functional significance of the conserved aromatic residues that line the narrow gorge leading to the buried active site, including recognition of the important roles played by some of them in π-cation and π-π interactions with substrates and inhibitors, in particular of the quaternary group of acetylcholine (ACh) with the indole rings of two conserved tryptophan residues. They also investigated the catalytic machinery of the enzyme, the functional and structural roles of conserved water molecules in binding and catalysis, and the functional significance of the high dipole moment aligned along the axis of the active-site gorge, which generates a large potential gradient pulling ACh towards the active site. The findings of Sussman and Silman have provide important input for the design and synthesis of anti-AChE drugs that are used in the management of neurodegenerative and neuromuscular diseases, such as Alzheimer’s and myasthenia gravis, for the development of improved antidotes and bioscavengers for the treatment of nerve agent intoxication, and for design of novel anticholinesterase insecticides. These latter studies were facilitated by their subsequent solution, in 2000, of the crystal structures of human AChE, and of that of Drosophila melanogaster AChE, the only insect AChE structure solved to date.

In recent years, Silman and Sussman have also collaborated with colleagues in the Department of Biomolecular Sciences Chemistry at the Weizmann, and extended their interests to structural studies of two other enzymes associated with degenerative diseases. With Prof. Tony Futerman, they were able to crystallize and determine the 3D structure of acid-β-glucosidase, the enzyme whose malfunction, due to mutation, results in Gaucher disease. The structural and functional data so generated have already resulted in novel approaches to the treatment of the disease. With Prof. Dan Tawfik, they solved the 3D structures of the serum enzyme, paraoxonase (PON), and of bacterial phosphotriesterases (PTEs), both of which hydrolyze organophosphate-based pesticides and nerve agents, and thus comprise a new generation of catalytic bioscavengers. PON is of interest, however, not only to toxicologists, but also to cardiovascular clinicians working in the field of atherosclerosis, the major degenerative disease in ageing populations.

Currently, Profs. Silman and Sussman are both Emeritus Professors at the Weizmann Institute of Science.

This Special Issue of Molecules, in honor of Profs. Silman and Sussman, welcomes manuscripts describing original work, as well as review articles, on the structures, modes of action and biological roles of AChE and the butyrylcholinesterase (BChE). The Guest Editors will be pleased to accept and review manuscripts that address the topics listed below, but not restricted to them:

• Methodological advances in research on the ChEs, both experimental and theoretical
• 3D-Structural studies on native ChEs, their complexes and their conjugates
• Kinetics and mechanism of action of ChEs
• Inhibition by natural and synthetic ligands
• Reactivation of ChEs inhibited by nerve agents and insecticides
• SAR studies on drugs acting on the ChEs in the central and peripheral nervous systems
• Non-cholinergic functions of the ChEs

Prof. Dr. Anthony H. Futerman
Dr. Yacov Ashani
Dr. Gabi Amitai
Dr. Lev Weiner
Guest Editors

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• Acetylcholinesterase
• Butyrylcholinestersae
• 3D structure
• Inhibitors and reactivators
• Enzymic mechanism
• Biological role