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=='''A Physical Model of Acetylcholinesterase in Complex with Acetylcholine and Fasciculin-II'''==
=='''Physical Models of Acetylcholinesterase in Complex with Acetylcholine and the Green Mamba Snake Toxin, Fasciculin-II'''==
Students: Mary Acheampong. Daviana Dueno, Bobby Glover, Alafia Henry, Randol Mata, Marisa VanBrakle.
Students: Mary Acheampong. Daviana Dueno, Bobby Glover, Alafia Henry, Randol Mata, and Marisa VanBrakle, Hostos-Lincoln Academy.


Teacher: Allison Granberry
Teacher: Allison Granberry, Hostos-Lincoln Academy


Mentors: Joel Sussman, Weissman Institule of Science, and Lars Westblade, touro College of Pharmacy.
Mentors: Joel Sussman, Weissman Institule of Science, and Lars Westblade, Touro College of Pharmacy.


==='''Abstract'''===
==='''Introduction'''===


Acetylcholinesterase(AChE) is essential for the hydrolysis of the neurotransmitter acetylcholine(ACh) in cholinergic synapses. Irreversible inhibition of AChE can lead to increased levels of ACh and ultimately death. Conversely, suppressed levels of ACh may lead to memory deficits associated with Alzheimer's disease. AChE has a deep(20A) and narrow(5A) gorge lined with 14 aromatic residues, with its active site at the bottom of the gorge. Initially, ACh binds to the peripheral anionic site(PAS)  of AChE and is funneled down the gorge to the active site by interactions between the aromatic rings of the 14 aromatic residues and the quaternary ammonium ion of ACh. At the active site, ACh is oriented for hydrolysis by interactions between the catalytic anionic ion site and the quaternary ammonium ion of ACh. The Fasciculin-II(FAS-II)toxin, from the East African Green Mamba snake(''Dendroaspis angusticeps'') venom, inhibits AChE by binding to the top of the active-site gorge, and thus preventing ACh from entering into it. The Hostos-Lincoln Academy SMART(Students Modeling A Research Topic) team and MSOE have designed and made two physical models by three-dimensional(3D) printing technology: ''Torpedo californica''(''Tc'') AChE in complex with a modeled ACh ligand and ''Tc''AChE in complex with FAS-II.
Acetylcholinesterase(AChE) is essential for the hydrolysis of the neurotransmitter acetylcholine(ACh), and therefore the termination of the nerve impulse in cholinergic synapses. Irreversible inhibition of AChE can lead to increased levels of ACh in cholinergic synapses and ultimately death. Conversely, suppressed levels of ACh may lead to memory deficits associated with Alzheimer's disease. AChE has a deep(20A) and narrow(5A) gorge lined with 14 aromatic residues, with its active site at the bottom of the gorge. Initially, ACh binds to the peripheral anionic site(PAS)  of AChE and is funneled down the gorge to the active site by interactions between the aromatic rings of the 14 aromatic residues and the quaternary ammonium ion of ACh. At the active site, ACh is oriented for hydrolysis by interactions between the catalytic anionic ion site and the quaternary ammonium ion of ACh. The Fasciculin-II (FAS-II)toxin, a component of the East African Green Mamba snake(''Dendroaspis angusticeps'') venom, inhibits AChE by binding to the top of the active-site gorge, including residues that form the PAS; thus preventing ACh from entering the active-site gorge. The Hostos-Lincoln Academy Students Modeling A Research Topic(S.M.A.R.T) team and the Center for BioMolecular Modeling have designed and fabricated two physical models using a combination of computational molecular modeling and three-dimensional(3D) printing technology: ''Torpedo californica''(''Tc'') AChE in complex with a modeled ACh ligand and ''Tc''AChE in complex with FAS-II.


==='''Designing a Physical Model to Tell the Story of Acetylcholinesterase'''===
==='''Designing Physical Models to Tell the Story of Acetylcholinesterase'''===


Reflected in our design are two key concepts of AChE: the biochemistry of how the ACh overcomes the depth of the active site gorge before hydrolysis can occur, and how a toxin inhibits the substrate from finding the active site.Two physical models were designed and made by 3-dimensional printing technology: ''Torpedo californica (Tc)'' AChE in complex with a modeled ACh ligand, and ''Tc'' AChE in complex with FAS-II. Both models were based on protein data bank (PDB) files, and Rasmol computer modeling program. PDB files included PDB entry code 2ace for the ''Tc''AChE/ACh complex, and PDB entry code 1fss for the ''Tc'' AChE/FAS-II complex.
Reflected in our design are two key concepts of AChE biology: the mechanism by which AChE hydrolyses ACh (the substrate traffic story), and how the Green Mamba Snake toxin, FAS-II, inhibits the hydrolysis of ACh (the inhibition story). Two physical models were designed and fabricated using a combination of computational molecular modeling and 3D printing technology: ''Tc'' AChE in complex with a modeled ACh ligand, and ''Tc'' AChE in complex with FAS-II. Both models were designed using the respective protein data bank (PDB) files: 2ace for the ''Tc''AChE/ACh complex and 1fss for the''Tc''AChE'FAS-II complex, and Rasmol computer modeling program.  


<applet load='2ace' size='300' frame='true' align='left' scene='Sandbox_250/Ache_ach/1' caption='AChE/ACh'/>
<applet load='2ace' size='300' frame='true' align='left' scene='Sandbox_250/Ache_ach/1' caption='AChE/ACh'/>




===='''Features of the Substrate Traffic Story: AChE/ACh'''====
===='''Features of the Substrate Traffic Story:''a Model of'' AChE/ACh'''====


<scene name='Sandbox_250/Ache_ach/5'>AChE</scene> is an alpha/beta hydrolase fold with an amino acid sequence of 4-535. <scene name='Sandbox_250/Ache_ach/6'>ACh</scene> consists of an acytoxy group, ethylene group and a quaternary ammonium ion.
The ''Tc''<scene name='Sandbox_250/Ache_ach/5'>AChE</scene> protein contains 537 amino acids and forms and is an α/β hydrolase fold. The neurotransmitter <scene name='Sandbox_250/Ache_ach/6'>ACh</scene> consists of an acytoxy group, an ethylene group and a positively charged quaternary ammonium ion.


The <scene name='Sandbox_250/Ache_ach/17'>14 aromatic residues</scene> that line the active site gorge are tyr70, trp84, trp120, tyr121, tyr130, trp233, trp279, phe288, phe290, phe330, phe331, tyr334, trp432 and tyr442.
The <scene name='Sandbox_250/Ache_ach/17'>14 aromatic residues</scene> that line the active site gorge are Tyr70, Trp84, Trp120, Tyr121, Tyr130, Trp233, Trp279, Phe288, Phe290, Phe330, Phe331, Tyr334, Trp432 and Tyr442. These aromatic residues interact with the positively charged quaternary ammonium ion of ACh by virtue of cation-π interactions to filter it down the active-site gorge to the catalytic triad.


The Peripheral Anionic Site(PAS) includes <scene name='Sandbox_250/Ache_ach/11'>Tyr279, Tyr70 and Tyr121</scene>. Initially, the positively charged quaternary ammonium ion of ACh is attracted to and binds to the <scene name='Sandbox_250/Ache_ach/12'>PAS of AChE</scene>, highlighted in yellow.  
The PAS includes residues <scene name='Sandbox_250/Ache_ach/11'>Tyr70, Tyr121 and Trp279</scene>. Initially, the positively charged quaternary ammonium ion of ACh is attracted to and binds to the <scene name='Sandbox_250/Ache_ach/12'>PAS of AChE</scene>, highlighted in yellow.  


The Catalytic Anionic Site(CAS) includes <scene name='Sandbox_250/Ache_ach/18'>Trp84 and Phe330</scene>. The  
The Catalytic Anionic Site (CAS) includes residues <scene name='Sandbox_250/Ache_ach/18'>Trp84 and Phe330</scene>. The  
<scene name='Sandbox_250/Ache_ach/14'>CAS</scene>, highlighted in red, holds ACh in the optimal position for hydrolysis by interacting with the quaternary ammonium ion of ACh.
<scene name='Sandbox_250/Ache_ach/14'>CAS</scene>, highlighted in red, holds ACh in the optimal position for hydrolysis by interacting with the quaternary ammonium ion of ACh.


The active site includes three residues: <scene name='Sandbox_250/Ache_ach/20'>Ser200, Glu327, and His440</scene>. The <scene name='Sandbox_250/Ache_ach/16'>Catalytic Triad</scene>, highlithed in blue, is responsible for the hydrolysis of ACh into acetate and choline.
The AChE active site includes three residues that form a catalytic triad: <scene name='Sandbox_250/Ache_ach/20'>Ser200, Glu327, and His440</scene>. The <scene name='Sandbox_250/Ache_ach/16'>Catalytic Triad</scene>, highlithed in blue, is responsible for the hydrolysis of ACh into acetate and choline.




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===='''Features of the Inhibition Story'''====
===='''Features of the Inhibition Story: a Model of AChE/FAS-II'''====


<scene name='Sandbox_250/Ache_fas2/9'>FAS-II</scene> is a 61-residue polypeptide with 4 beta sheets forming three loops or fingers.   
The Green Mamba snake toxin, <scene name='Sandbox_250/Ache_fas2/9'>FAS-II</scene>, is a 61-residue protein that folds into 4β sheets forming 3β sheets forming loops or fingers.   


FAS-II is attracted to AChE by a number of mechanisms:
FAS-II binds to and inhibits AChE using two major mechanisms:
 
1. Amino acid specificity: residues <scene name='Sandbox_250/Ache_fas2/14'>Thr8, Arg27 and Met33</scene> are located on two of the three fingers of FAS-II. When FAS-II <scene name='Sandbox_250/Ache_fas2/12'>binds</scene> to AChE, Arg27 and Met33 interact with Trp279 part of the PAS, while Thr8 interact with Tyr70, also part of the PAS.
 
2. Shape: Once bound to the PAS, two loops of FAS-II fit in to the AChE active-site gorge like a hand fits into a glove. Once this occurs, the entrance of the gorge is <scene name='Sandbox_250/Ache_fas2/13'>blocked</scene> such that acetylcholine may not enter, and therefore it will not be hydrolysed. This results in the increased levels of AChE in the cholinergic synapse, and ultimately death.


1. Amino acid specificity: <scene name='Sandbox_250/Ache_fas2/14'>Thr8, Met33 and Arg27</scene> are located on two of the three fingers of the FAS-II. When FAS-II <scene name='Sandbox_250/Ache_fas2/12'>binds</scene> to AChE, Arg27 and Met33 interact with Trp279 of the Peripheral Anionic Site, while Thr8 interact with Tyr70 of the Peripheral Anionic Site.


2. Shape: Once bound to the PAS, two loops of FAS-II fit in to the active-site gorge like a hand fits into a glove. Once this interaction occurs, the entrance of the gorge is <scene name='Sandbox_250/Ache_fas2/13'>blocked</scene> such that acetylcholine may not enter.




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==='''Acknowledgements'''===
==='''Acknowledgements'''===


1. Bill & Melinda Gates Foundation
1. Howard Hughes Medical Institue Pre-College Program
2. Center for BioMolecular  Modeling, Milwaukee School of Engineering


2. Howard Hughes Medical Institue Pre-College Program
3. The Rockefeller University Center for Clinical and Translational Science
 
3. Center for BioMolecular  Modeling, Milwaukee School of Engineering
4. The Rockefeller University S.M.A.R.T Team Program


4. The Rockefeller University Center for Clinical and Translational Science
5. The Rockefeller University Science Outreach Program


5. The Rockefeller University SMART Team Program
6. Touro College of Pharmacy


6. The Rockefeller University Science Outreach Program
7. Michal Harel, Weizmann Institute of Science


7. Malcolm Twist
8. Malcolm Twist

Proteopedia Page Contributors and Editors (what is this?)Proteopedia Page Contributors and Editors (what is this?)

Joel L. Sussman, Allison Granberry, Jaime Prilusky
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