Engineering a Highly Regioselective Fungal Peroxygenase for the Synthesis of Hydroxy Fatty Acids

Autor: Patricia Gomez de Santos, Alejandro González‐Benjumea, Angela Fernandez‐Garcia, Carmen Aranda, Yinqi Wu, Andrada But, Patricia Molina‐Espeja, Diana M. Maté, David Gonzalez‐Perez, Wuyuan Zhang, Jan Kiebist, Katrin Scheibner, Martin Hofrichter, Katarzyna Świderek, Vicent Moliner, Julia Sanz‐Aparicio, Frank Hollmann, Ana Gutiérrez, Miguel Alcalde
Přispěvatelé: European Commission, Ministerio de Ciencia e Innovación (España), Agencia Estatal de Investigación (España), Comunidad de Madrid, Ministerio de Ciencia, Innovación y Universidades (España), Gómez de Santos, Patricia, González-Benjumea, Alejandro, Aranda, Carmen, Wu, Yinqi, Molina-Espeja, Patricia, Maté, Diana M., González-Pérez, David, Zhang, Wuyuan, Kiebist, Jan, Hofrichter, Martin, Świderek, Katarzyna, Moliner, Vicent, Sanz Aparicio, Julia, Hollmann, Frank, Gutiérrez Suárez, Ana, Alcalde Galeote, Miguel
Rok vydání: 2023
Předmět:
Zdroj: Angewandte Chemie (International Edition), 62(9)
ISSN: 1521-3773
1433-7851
Popis: 10 páginas.- 4 figuras.- 2 tablas.- 30 referencias.- Supporting information for this article is given via a link at the end of the document https://onlinelibrary.wiley.com/doi/10.1002/anie.202217372
he hydroxylation of fatty acids is an appealing reaction in synthetic chemistry, although the lack of selective catalysts hampers its industrial implementation. Here, we have engineered a highly regioselective fungal peroxygenase for the w-1 hydroxylation of fatty acids with quenched stepwise over-oxidation. One single mutation near the Phe catalytic tripod narrowed the heme cavity, promoting a dramatic shift toward sub-terminal hydroxylation with a drop in the over-oxidation activity. While crystallographic soaking experiments and molecular dynamic simulations shed light on this unique oxidation pattern, the selective biocatalyst was produced by Pichia pastoris at 0.4 g/L in a fed-batch bioreactor and used in the preparative synthesis of 1.4 g of (w-1)-hydroxytetradecanoic acid with 95% regioselectivity and 83% ee through the (S)-enantiomer.The hydroxylation of fatty acids is an appealing reaction in synthetic chemistry, although the lack of selective catalysts hampers its industrial implementation. Here, we have engineered a highly regioselective fungal peroxygenase for the w-1 hydroxylation of fatty acids with quenched stepwise over-oxidation. One single mutation near the Phe catalytic tripod narrowed the heme cavity, promoting a dramatic shift toward sub-terminal hydroxylation with a drop in the over-oxidation activity. While crystallographic soaking experiments and molecular dynamic simulations shed light on this unique oxidation pattern, the selective biocatalyst was produced by Pichia pastoris at 0.4 g/L in a fed-batch bioreactor and used in the preparative synthesis of 1.4 g of (w-1)-hydroxytetradecanoic acid with 95% regioselectivity and 83% ee through the (S)-enantiomer.
This work was supported by the European Union Project grant H2020-BBI-PPP-2015-2-720297-ENZOX2, the I+D+I PID2019-106166RB-I00-OXYWAVE Spanish project funded by the Ministerio de Ciencia e Innovación/Agencia Estatal de Investigación (AEI)/doi: 10.13039/501100011033/, the PID2020-118968RB-I00 LILI project from the Spanish MCIN/AEI/10.13039/501100011033, the ‘Comunidad de Madrid’ Synergy CAM project Y2018/BIO-4738-EVOCHIMERA-CMand the PIE-CSIC projects PIE-202040E185 and PIE-201580E042. PGS thanks the Ministry of Science, Innovation and Universities (Spain) for her FPI scholarship (BES-2017-080040) and to the Ministry of Science and Innovation for her contract as part of PTQ2020-011037 project funded by MCIN/AEI/10.13039/501100011033 within the NextGenerationEU/PRTR.DGP thanks Juan de la Cierva Incorporación contract Ref nº: IJC2020-043725-I, funded by MCIN / AEI / 10.13039/501100011033, and the EUNextGenerationEU/PRTR program. The authors thank the Synchrotron Radiation Source at Alba (Barcelona, Spain) for assistance with the BL13-XALOC beamline.
Databáze: OpenAIRE
Externí odkaz: