AChE and Inhibition

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You may include any references to papers as in: the use of JSmol in Proteopedia ^[1] or to the article describing Jmol ^[2] to the rescue.

History

In order to further understand acetylcholinesterase and its medical implications, it is important to look at the discovery of its substrate, acetylcholine. Although acetylcholine had been previously identified and studied, it wasn’t until 1914 when Sir Henry Hallet Dale discovered its function as a chemical mediator. Dale was able to determine that ACh was necessary to transmit certain electrical impulses within the human body. His findings were later confirmed by Otto Loewi and this was celebrated as the first identification of a neurotransmitter. Both Dale and Loewi were awarded the Nobel Prize in Physiology and Medicine in 1936 for their findings. ^[3] It wasn’t until years later that further studies were performed to discern more fully the function of acetylcholine in the body and specifically how it is recycled. It was during this time that acetylcholinesterase was realized and its role in the function of ACh more fully understood. In more recent years, more extensive work has been completed to look at the medical implications of acetylcholinesterase and more importantly how inhibitors of it could be used as symptomatic treatment in some diseases. ^[3]

Function

Acetylcholinesterase plays an essential role in neurotransmission throughout the human body, including the central nervous system and muscular systems. Specifically, its function is in relation to the neurotransmitter acetylcholine. ^[4] Acetylcholine, synthesized in nerve terminals from acetyl CoA and choline, works at cholinergic synapses within the nervous system. While some neurotransmitters are terminated at the postsynaptic terminal by reuptake, ACh is broken down by acetylcholinesterase through the process of hydrolysis. ^[5] This hydrolysis breaks acetylcholine down into an acetate and choline. AChE has a relatively high catalytic activity; each molecule of AChE can degrade 25000 molecules of ACh per second. ^[6] Later, these are recycled to again form acetylcholine for use in these same neuromuscular junctions throughout the body or within the central nervous system itself. Importance of the enzyme AChE is shown during inhibition with a nerve gas such as sarin. With this irreversible inhibitor present, there is an excess of acetylcholine within the synapses and continued activation of the ACh receptors. This can have a number of dire effects such as respiratory and/or cardiac dysfunction and possible death. However, reversible acetylcholinesterase inhibitors can be used in numerous therapeutic medications to treat symptoms related to diseases such as Alzheimer’s disease and myasthenia gravis. ^[5]

Structure

Acetylcholinesterase is a serine hydrolase, described to have an ellipsoidal shape. A monomer of the enzyme contains 12 mixed β sheet surrounded by 14 α helices. The enzyme does contain one somewhat remarkable feature, a deep and narrow gorge which is ~ 20 Ǻ long penetrating halfway into the enzyme. The active site within AChE is composed of two subsites, an anionic subsite and an esteratic subsite; both subsites are important in the function of the enzyme. Contained within the narrow gorge leading to the active site, there are 14 aromatic residues. These residues have been highly conserved across all species who rely on acetylcholinesterase, and therefore are known to be essential in its processes. There is an additional peripheral binding site within AChE which is distinct from the primary acetylcholine binding site. This serves as a binding site for uncompetitive inhibitors and is clearly separate from the site occupied by competitive inhibitors. Therefore, this is the site thoroughly studied by pharmaceutical companies developing AChE inhibitors as a treatment for a number of diseases (Colović et al 2013).

Synthesis and Localization

While the enzyme acetylcholinesterase may be slightly adjusted from organism to organism, each maintains the essential residues to carry out its functions. The synthesis of this enzyme has been studied over the years and a number of mechanisms and interactions have emerged. Synthesis of AChE occurs on the rough endoplasmic reticulum. There, it is assembled into dimers and tetramers and later reassembled into collagen-tailed molecules (Rotundo et al, 2008). While a number of these molecules are not catalytically active and eventually are broken down, those that mature into catalytically active enzyme are transported to and through the Golgi apparatus. AChE is then transferred to the cell surface where it briefly interacts with the extracellular matrix before later being covalently attached. It has been determined that AChE localizes at the neuromuscular junction through its interactions with perlecan, a proteoglycan. Additionally, the carboxyl terminal domain of the collagen-like tail is necessary for this attachment and localization of AChE to the synapse of neuromuscular junctions (Rotundo et al, 2008).

Reactions

We have already discussed the main elements of the structure of acetylcholinesterase; it is this structure and the localization of the pockets within it that contribute to its reactability. Within the number of binding sites within the enzyme, there is a catalytic triad of amino acids: serine 200, histidine 440, and glutamate 327, similar to other serine hydrolases. It is also known that the residues Tryptophan 84 and Phenylalanine 330 are important in the ligand recognition. Following the hydrolysis of acetylcholine, in which ACh binds directly to the serine 200, there is the formation of an acyl-enzyme and free choline. This enzyme then undergoes nucleophilic attach by a water molecule and assisted by a histidine 440 group. This frees the acetic acid from the active site and regenerates the free enzyme, allowing it to bind other acetylcholine molecules (Colović et al, 2013). The process breaks down acetylcholine into an acetate and choline, thereby terminating the neurotransmission.

Regulation

As with all processes throughout the human body, it is important to be able to regulate the activity of acetylcholinesterase within both the nervous and musculoskeletal systems. AChE activity is known to be related to skeletal muscle use. It has been shown that by inhibiting muscle contraction, activity of AChE is decreased (Fernandez-Valle & Rotundo, 1998). Additionally, further studies have been completed to describe the role of chaperones in the assembly of AChE and how these are used to regulate the amount of acetylcholinesterase within the body. There are a number of molecular chaperones used to aid in the correct folding and assembly of AChE after synthesis. Studies have shown that overexpression of chicken endoplasmic reticulum chaperones ERP72 and protein disulfide isomerase (PDI) led to increased expression and activity of junctional AChE (Rotundo et al, 2008). Conversly, it was shown that with inhibition of these chaperones, such as PDI, there was a decrease in the expression of intracellular AChE. Another process involved in the regulation of AChE occurs in translational control by the mRNA-binding protein Pumilio2. It has been shown that overexpression of PUM2 represses AChE translation, while knockout of this transcript increases expression. It is thought that PUM2 could be involved both in the localization of AChE and the translational control mechanism throughout activity-dependent de-repression (Rotundo et al., 2008).

Medical Implications

As the scientific community has continued to advance their knowledge of acetylcholinesterase and its function, a number of medical advances have been made as well. As AChE plays such a major role in the function of our nervous systems, primarily in the hydrolysis of acetylcholine to acetate and choline, numerous therapeutics have been developed to oppose this function (Colović et al, 2013). These acetylcholinesterase inhibitors are able to prevent this hydrolysis and therefore maintain higher levels of acetylcholine and longer duration of action within the synapses of neuromuscular junctions and cholinergic brain synapses. It is important to note that AChE inhibitors can be broken into two groups: reversible and irreversible. Irreversible inhibitors have a number of toxic effects and are occasionally used as insecticides. One of the main uses of reversible acetylcholinesterase inhibitors is in the treatment of Alzheimer’s disease. The primary characteristic of AD is dementia, described as memory loss and other intellectual abilities which interfere with daily life. Primarily, the cause of this disease is in the loss of brain cholinergic neurons and a decrease of the neurotransmitter acetylcholine. While AChE inhibitors due not cure Alzheimer’s, they are frequently used in the treatment of the symptoms related to memory, thinking, and judgement. Acetylcholinesterase inhibitors can also be used in the treatment of diseases such as myasthenia gravis, glaucoma, and as an antidote to anticholinergic overdose.

References

1. ↑ Hanson, R. M., Prilusky, J., Renjian, Z., Nakane, T. and Sussman, J. L. (2013), JSmol and the Next-Generation Web-Based Representation of 3D Molecular Structure as Applied to Proteopedia. Isr. J. Chem., 53:207-216. doi:http://dx.doi.org/10.1002/ijch.201300024
2. ↑ Herraez A. Biomolecules in the computer: Jmol to the rescue. Biochem Mol Biol Educ. 2006 Jul;34(4):255-61. doi: 10.1002/bmb.2006.494034042644. PMID:21638687 doi:10.1002/bmb.2006.494034042644
3. ↑ ^{3.0 3.1} Jacob, L. M. (2018). Acetylcholine. Salem Press Encyclopedia of Science. Retrieved from http://proxy.library.maryville.edu/login?url=https://search.ebscohost.com/login.aspx?direct=true&db=ers&AN=87322209&site=eds-live&scope=site
4. ↑ Downes, G. B., & Granto, M. (2004). Acetylcholinesterase function is dispensable for sensory neurite growth but is critical for neuromuscular synapse stability. In Developmental Biology (1st ed., Vol. 280, pp. 232-245). doi:https://doi.org/10.1016/j.ydbio.2004.02.027
5. ↑ ^{5.0 5.1} Purves D, Augustine GJ, Fitzpatrick D, et al., editors. Neuroscience. 2nd edition. Sunderland (MA): Sinauer Associates; 2001. Acetylcholine.Available from: https://www.ncbi.nlm.nih.gov/books/NBK11143/
6. ↑ Colović, M. B., Krstić, D. Z., Lazarević-Pašti, T. D., Bondžić, A. M., & Vasić, V. M. (2013). Acetylcholinesterase inhibitors: pharmacology and toxicology. Current neuropharmacology, 11(3), 315-35.