Acetylcholinesterase is a serine hydrolase that belongs to the esterase family within higher eukaryotes. This family acts on different types of carboxylic esters. Acetylcholinesterase's biological role is the termination of impulse transmissions at cholinergic synapses within the nervous system by rapid hydrolysis of the neurotransmitter, acetylcholine
Acetylcholinesterase is an ellipsoidal molecule approximately 45 x 60 x 65 angstroms, which consists of a 12 stranded central mixed beta sheet surrounded by 14 alpha helices (Sussman et al, 1991). Studies have indicated several major domains within the protein: a catalytic active site composed of two subsites, the aromatic gorge in which the catalytic active site lies, and a peripheral anionic site, distinct from the catalytic active site, which plays a role in the confirmation of the residues within the aromatic gorge and active site. This view of the protein is provided for easier viewing of the 3-D structure of the entire protein in stereo.
The active site is composed of two subsites: the esteratic subsite which contains the catalytic triad, and the anionic subsite that accommodates the positive quaternary pole of acetylcholine. The esteratic subsite contains the catalytic machinery of the enzyme: a catalytic triad of Ser 200, His 440, and Glu 327. This catalytic triad is similar to other serine proteases, except that this triad is the first to show Glu as the third member as opposed to Asp. In addition, the triad is of opposite handedness to that of the other proteases. The anionic subsite is defined by Trp 84, Phe 330, and Phe 331. Its role is to orient the charged part of the substrate that enters the active center. This role is the main function of the Trp residue (Sussman et al, 1991). This subsite has another interesting characteristic; it is involved in a "cross-talk" mechanism with the peripheral anionic site which will be discussed later.
The aromatic gorge in the protein is approximately 20 angstroms deep and penetrates halfway into the enzyme. The active site lies at the base of this gorge only 4 angstroms above the base, leading some to label this the active gorge. The aromatic gorge is a more appropriate term, though, because 40% of its lining is composed of 14 aromatic residues which are highly conserved from different species of acetylcholinesterases (Harel et al, 1993). The high aromatic content of the walls and floor may explain why studies have proposed hydrophobic and anionic binding sites independent of the active site. Only a few acidic residues are present within the gorge.
The most interesting aspect of this enzyme is the peripheral anionic site (PAS) on its surface. Site-directed labeling and mutagenesis studies place the location of the PAS at or near the rim of the aromatic gorge (Barak et al, 1994). This site has the ability to bind to many different types of ligands, and by doing so effects the conformation of the active center. Six residues have established activity within this site: Trp 286, Tyr 72, Tyr 124, Glu 285, and Asp 74 and Tyr 341, which are located on the opposite side of the gorge entrance to the previous four. This array of residues exhibits flexibility which accommodates many distinct ligands, and also implies their conformational mobility (Ordentlich et al, 1993). The common feature of these conformations is a core comprised of Trp 286 and Asp 74.
As mentioned above, the peripheral anionic site is involved in a "cross-talk" mechanism with the active site. This mechanism is an interaction between the Trp 286 and Trp 86 residues. When Trp 286 is bonded on the periphery, it effects the Trp 86 in the active site and causes distinct conformations of the active site to occur, thus changing the functionality of the enzyme. Asp 74 also plays a role in allosteric modulation of the enzyme. This residue acts with Tyr 341, as well as other residues along the aromatic gorge, and terminates at Tyr 337. The sensitivity of these residues and the plasticity of the active center are probably the result of evolutionary design aimed to confer optimal catalytic activity under a wide variety of conditions that are characteristic for the operation of acetylcholinesterase in the synaptic cleft.
