3D structure of acetylcholinesterase
Key Enzyme in the Nervous System
Acetylcholinesterase (AChE) is key enzyme in the cholinergic pathways in the animal nervous system. By rapid hydrolysis of the neurotransmitter, acetylcholine (ACh), AChE terminates neurotransmission at cholinergic synapses. AChE is a very fast enzyme, especially for a serine hydrolase, functioning at a rate approaching that of a diffusion-controlled reaction. The powerful toxicity of organophosphorus (OP) poisons is attributed primarily to their potent AChE inhibitors. AChE inhibitors are utilized in the treatment of various neurological and are the key drugs approved by the FDA for management of Alzheimer's disease (AD). Many carbamates and OPs serve as potent insecticides, by selectively inhibiting insect AChE.
In 1991, at the Weizmann Institute, Joel Sussman & Israel Silman, and thier colleagues, determined the 3D structure of AChE from Torpedo californica (TcAChE), permitting visualization, for the first time, at atomic resolution, of a binding pocket for ACh. It also allowed identification of the active site of AChE, which, unexpectedly, is located at the bottom of a , lined largely by aromatic residues. This unusual structure permitted them to work out structure-function relationships for AChE. The so-called 'anionic' binding site for the quaternary moiety of ACh does not contain several negative charges, as earlier postulated. However, AChE shows a remarkable asymmetric charge distribution resulting in an unusually large dipole moment (~1,700 Debye) aligned along the active-site gorge. Modeling studies suggested that the quaternary group interacts primarily with the indole ring of the conserved tryptophan residue, W84, via cation-π interaction, as well as with F330. Crystallographic studies on several AChE-ligand complexes confirmed this. This was in agreement with labeling studies in solution and theoretical studies on the cation-π interaction. From the various inhibitor/AChE complexes they have studied, including most currently available and potential drugs for treatment of AD, they see that although many interact very tightly with AChE (binding constants 1010), interaction is mediated mostly via waters, and van der Waals interactions, with few direct interactions with the protein.
The increasing longevity of people's lifespans, and the resulting increased
prevelance of dementias such as Alzheimers Disease, led scientists to study the animal enzyme Acetylcholinesterase (AChE) as a possible cause. This enzyme rapidly degrades or hydrolizes the neurotransmitter acetylcholine in synapses (junctions between nerve cells) of cholinergic nerve pathways into acetic acid and choline, to turn off the chemical signal for the nerve to fire. Should something happen to deactivate or kill this vital enzyme, nervous paralysis of vital functions occurs, leading to rapid death. Although AChE is apparently not the cause of Alzheimers, it does seem to play a minor role, in that weak inibitory drugs such as Tacrine, E2020 (Aricept) and the natural Chinese natural produce Huperzine appear to delay symptoms. Furthermore, this enzyme is a key target of some very important nerve gasses and related insecticides. Furthermore, it is a pretty fascinating enzyme to study.
Because of the relative ease in obtaining purified protein in abundance, Aceytlcholinesterase was first crystallized by Joel Sussman of the Weizmann Institute, Rehovot Israel, after being extracted from the electric organ of the Pacific Sting Ray, Torpedo californica, order to determine its detailed
3-dimensional structure by X-ray crystallography (2ace). Subsequently its X-ray structure has been determined from over 20 species, ranging from the fruit fly Drosophila to human.
Acetylcholinesterase is a fairly large protein, consisting of at least 535 amino acid residues in a single peptide chain, that folds into a single protein domain without any apparent symmetry.
The active site region of this enzyme has two sites, a catalytic site and a peripheral site, which helps prebind the substrate and direct it toward the active site. When the 3-D structure was first determined, the big surprise was
that the active site was deep inside the protein, at the end or base of a
,
lined with aromatic residues, with the peripheral site at the top or lip of this gorge. Amazingly, there were no acidic or negatively charged
residues anywhere in these 2 sites or along this gorge, as would be expected to
help attract and bind the basic, positively charged acetylcholine substrate, although are are some acidic residues nearby. Instead, bulky aromatic residues (These numbers are the sequential numbering
of the residues, starting from the N-terminus, according to the Torpedo Californica form of the enzyme.)
AChE inhibitors and substrates
Among the most interesting drugs that have been designed to inhibit this enzyme are those that
have two binding sites that bind both the peripheral and catatylic sites simultaneously. Such
drugs bind highly specificly and strongly. A good example is
.
It appears that the principal interaction between the aceylcholine and the enzyme is relatively newly discovered cation-pi interactions between the cationic moiety of the substrate and the many aromatic residues lining the catalytic gorge. Unlike most
interatomic interactions in chemistry, cation-pi interactions are unusual in that their energy hardly changes as the cationic and
aromatic ring centers vary between 4 and 7 Angstroms apart, and for a wide variety of relative orientations of the aromatic rings.
This gives the substrate an energetically smooth ride down the gorge with few bumps or barriers to impede passage down the gorge.
Most acetylcholinesterases have a net negative charge and a large patch of negative potential around the entrance to the active site gorge, which may be useful to attract the positively charged acetycholine substrate to the site. As one travels down the gorge, this potential becomes increasingly more and more negative, reaching a peak at the active site at the base. Because of this potential, the peripherial site is thought to act like a substrate trap, that forces practically molecule of substrate that reaches
the peripheral site to travel down the gorge to the active site, that probably contributes greatly to the extremely rapid rate of degrading the substrate. This whole enzyme therefore acts like a brilliantly designed natural vacuum cleaner that clears the
neurotransmitter out of the synapse extremely quickly. Yet to be solved, however, is how the products clear the active site rapidly,
whether back through the gorge, or out a back door on the other side of the protein that quickly opens each catalytic cycle (Try 84
is actually near the surface of the 'underside' of the protein.)
Selected PDB 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.
- 1vzj Model complex of the Cholinesterase tetramer.
More structures can be obtained by searching for 'AChE' at the left <-.
