Sandbox Reserved 1852: Difference between revisions
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The Diels Alderase aims to catalyze the Diels-Alder reaction between 4-carboxybenzyl-trans-1,3-butadiene-1-carbamate and N,N-dimethylacrylamide for use in synthetic organic chemistry. Specifically, the enzyme surpasses uncatalyzed reactions by generating a product that is entirely [https://en.wikipedia.org/wiki/Stereoselectivity#:~:text=In%20chemistry%2C%20stereoselectivity%20is%20the,of%20a%20pre%2Dexisting%20one. stereoselective] for the 3R,4S endo form. The Diels-Alderase was built using ''de novo'' enzyme design, using computational modeling and refinement through collaborative problem-solving from online users. The first generation Diels-Alderase was made using the [https://en.wikipedia.org/wiki/Rosetta@home Rosetta] computational design program, where a potential active site was built and tested against a library of scaffold proteins. Later, as the active site was perfected, future generations of the Diels-Alderase were made using an online protein folding game called [https://en.wikipedia.org/wiki/Foldit Foldit,] where players competed to improve binding efficiency by completing various challenges.<ref name="Eiben"/> | The Diels Alderase aims to catalyze the Diels-Alder reaction between 4-carboxybenzyl-trans-1,3-butadiene-1-carbamate and N,N-dimethylacrylamide for use in synthetic organic chemistry. Specifically, the enzyme surpasses uncatalyzed reactions by generating a product that is entirely [https://en.wikipedia.org/wiki/Stereoselectivity#:~:text=In%20chemistry%2C%20stereoselectivity%20is%20the,of%20a%20pre%2Dexisting%20one. stereoselective] for the 3R,4S endo form. The Diels-Alderase was built using ''de novo'' enzyme design, using computational modeling and refinement through collaborative problem-solving from online users. The first generation Diels-Alderase was made using the [https://en.wikipedia.org/wiki/Rosetta@home Rosetta] computational design program, where a potential active site was built and tested against a library of scaffold proteins. Later, as the active site was perfected, future generations of the Diels-Alderase were made using an online protein folding game called [https://en.wikipedia.org/wiki/Foldit Foldit,] where players competed to improve binding efficiency by completing various challenges.<ref name="Eiben"/> | ||
[[Image: | [[Image:Dielsalderasesubstrates.jpeg|350px|left|thumb|Figure 1. Diels-Alderase substrates. Diene is 4-carboxybenzyl trans-1,3-butadiene-1-carbamate; dienophile is N,N- dimethylacrylamide.]] | ||
The Diels-Alderase was designed to connect a diene and dienophile in a [https://en.wikipedia.org/wiki/Diels%E2%80%93Alder_reaction Diels-Alder reaction.] It accomplishes this by decreasing the energy gap between the dienophile’s lowest unoccupied molecular orbital [https://en.wikipedia.org/wiki/HOMO_and_LUMO (LUMO)] and the diene’s highest occupied molecular orbital [https://en.wikipedia.org/wiki/HOMO_and_LUMO (HOMO)] in the transition state.<ref name="Siegel">PMID:20647463</ref> The current most active form of the Diels-Alderase is modelled under the PDB code [https://www.rcsb.org/structure/4O5T 4o5t]. | The Diels-Alderase was designed to connect a diene and dienophile in a [https://en.wikipedia.org/wiki/Diels%E2%80%93Alder_reaction Diels-Alder reaction.] It accomplishes this by decreasing the energy gap between the dienophile’s lowest unoccupied molecular orbital [https://en.wikipedia.org/wiki/HOMO_and_LUMO (LUMO)] and the diene’s highest occupied molecular orbital [https://en.wikipedia.org/wiki/HOMO_and_LUMO (HOMO)] in the transition state.<ref name="Siegel">PMID:20647463</ref> The current most active form of the Diels-Alderase is modelled under the PDB code [https://www.rcsb.org/structure/4O5T 4o5t]. | ||
[[Image:DAstereoselectivity.png|350px|left|thumb|Figure.x]] | |||
[[Image:SampleDAmech.jpeg|350px|right|thumb|Figure X.]] | |||
The binding pocket of 4o5t is selective for two substrates, 4-carboxybenzyl trans-1,3-butadiene-1-carbamate ([https://en.wikipedia.org/wiki/Diene diene]) and N,N- dimethylacrylamide (dienophile). These substrates are shown as a single, combined ligand–name—in the protein model. The binding site contains a [https://en.wikipedia.org/wiki/Hydrogen_bond hydrogen bond] donor (Tyr134) which lowers the LUMO energy and stabilizes the negative charge on the dienophile and a hydrogen bond acceptor (Glu208) that increases the HOMO energy and stabilizes the positive charge on the diene.<ref name="Siegel"/> Both of these H-bonding interactions work to stabilize the transition state, while also orienting the substrates in optimal conformations for reacting. | The binding pocket of 4o5t is selective for two substrates, 4-carboxybenzyl trans-1,3-butadiene-1-carbamate ([https://en.wikipedia.org/wiki/Diene diene]) and N,N- dimethylacrylamide (dienophile). These substrates are shown as a single, combined ligand–name—in the protein model. The binding site contains a [https://en.wikipedia.org/wiki/Hydrogen_bond hydrogen bond] donor (Tyr134) which lowers the LUMO energy and stabilizes the negative charge on the dienophile and a hydrogen bond acceptor (Glu208) that increases the HOMO energy and stabilizes the positive charge on the diene.<ref name="Siegel"/> Both of these H-bonding interactions work to stabilize the transition state, while also orienting the substrates in optimal conformations for reacting. | ||
