Fatty-acid oxygenation by fungal peroxygenases: From computational simulations to preparative regio- and stereoselective epoxidation
Epoxidation of unsaturated fatty acids by unspecific peroxygenases (UPOs) of the best-known long-UPO subfamily, including the Agrocybe aegerita (AaeUPO) and Coprinopsis cinerea enzymes, is reported here. To understand the different oxygenation patterns by members of the long-UPO and short-UPO subfam...
| Autores: | , , , , , , , , , , |
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| Tipo de recurso: | artículo |
| Fecha de publicación: | 2020 |
| País: | España |
| Institución: | Universitat Politècnica de Catalunya (UPC) |
| Repositorio: | UPCommons. Portal del coneixement obert de la UPC |
| Idioma: | inglés |
| OAI Identifier: | oai:upcommons.upc.edu:2117/345962 |
| Acceso en línea: | https://hdl.handle.net/2117/345962 https://dx.doi.org/10.1021/acscatal.0c03165 |
| Access Level: | acceso abierto |
| Palabra clave: | Monte Carlo method Chemistry, Analytic Computational chemistry Unsaturated fatty acids Oxygenation patterns Regioselective epoxidation Stereoselective epoxidation Monte Carlo molecular simulations Adaptive-PELE Fungal unspecific peroxygenases Montecarlo, Mètode de Química analítica Àrees temàtiques de la UPC::Informàtica::Aplicacions de la informàtica Àrees temàtiques de la UPC::Física |
| Sumario: | Epoxidation of unsaturated fatty acids by unspecific peroxygenases (UPOs) of the best-known long-UPO subfamily, including the Agrocybe aegerita (AaeUPO) and Coprinopsis cinerea enzymes, is reported here. To understand the different oxygenation patterns by members of the long-UPO and short-UPO subfamilies, the latter represented by the Marasmius rotula enzyme (MroUPO), fatty-acid diffusion into their heme pockets was simulated with the adaptive PELE software. Computational results shed light on the inability of AaeUPO to epoxidize oleic acid (C18:1), opposed to MroUPO, due to steric hindrances to harbor (with a good interaction energy) the substrate with the ¿9 C10 atom at a catalytically relevant distance (<3.5 Å) from the oxo group in simulated heme compound-I. However, effective a-linolenic acid epoxidation is anticipated because the ¿15 C16 atom would attain such a distance in AaeUPO thanks to its more terminal position. The above hypothesis was verified using an engineered MroUPO variant (I153F/S156F) with a narrowed heme access channel mimicking that of AaeUPO. Experimental oxygenation of unsaturated fatty acids by this variant thus resembles that of AaeUPO, including regioselective (from 95% to >99%) formation of cis,cis-15,16-epoxyoctadeca-9,12-dienoic acid. The nearly complete conversion of a-linolenic acid by the two enzymes was transferred to a small preparative scale, the yield of purified product was estimated, its chemical structure analyzed by NMR, and more interestingly, stereoselective production of the 15(R),16(S) enantiomer (80–83% ee) assessed by chiral HPLC. This enzymatic synthesis overcomes the unspecificity of chemical epoxidation where the reaction cannot be restricted to the formation of monoepoxides as found during m-perchlorobenzoic acid oxidation of a-linolenic acid. Moreover, the variant was able to produce the unsaturated dicarboxylic fatty acid, together with subterminal oxygenation products, during partial conversion of oleic acid. These two noteworthy reactions had not been reported for any UPO described to date. |
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