Rational Design of Resveratrol O-methyltransferase for the Production of Pinostilbene
Pinostilbene is a monomethyl ether analog of the well-known nutraceutical resveratrol. Both compounds have health-promoting properties, but the latter undergoes rapid metabolization and has low bioavailability. O-methylation improves the stability and bioavailability of resveratrol. In plants, these...
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doaj-0e5b392ab645468bad1d5fd5b6204af22021-04-21T23:07:23ZengMDPI AGInternational Journal of Molecular Sciences1661-65961422-00672021-04-01224345434510.3390/ijms22094345Rational Design of Resveratrol O-methyltransferase for the Production of PinostilbeneDaniela P. Herrera0Andrea M. Chánique1Ascensión Martínez-Márquez2Roque Bru-Martínez3Robert Kourist4Loreto P. Parra5Andreas Schüller6Department of Chemical and Bioprocesses Engineering, School of Engineering, Pontificia Universidad Católica de Chile, Vicuña Mackenna 4860, Santiago 7820244, ChileDepartment of Chemical and Bioprocesses Engineering, School of Engineering, Pontificia Universidad Católica de Chile, Vicuña Mackenna 4860, Santiago 7820244, ChileDepartment of Agrochemistry and Biochemistry, Faculty of Science and Multidisciplinary Institute for Environmental Studies “Ramon Margalef”, University of Alicante, 03690 Alicante, SpainDepartment of Agrochemistry and Biochemistry, Faculty of Science and Multidisciplinary Institute for Environmental Studies “Ramon Margalef”, University of Alicante, 03690 Alicante, SpainInstitute of Molecular Biotechnology, Graz University of Technology, Petersgasse 14, 8010 Graz, AustriaInstitute for Biological and Medical Engineering, Schools of Engineering, Medicine and Biological Sciences, Pontificia Universidad Católica de Chile, Vicuña Mackenna 4860, Santiago 7820244, ChileInstitute for Biological and Medical Engineering, Schools of Engineering, Medicine and Biological Sciences, Pontificia Universidad Católica de Chile, Vicuña Mackenna 4860, Santiago 7820244, ChilePinostilbene is a monomethyl ether analog of the well-known nutraceutical resveratrol. Both compounds have health-promoting properties, but the latter undergoes rapid metabolization and has low bioavailability. O-methylation improves the stability and bioavailability of resveratrol. In plants, these reactions are performed by O-methyltransferases (OMTs). Few efficient OMTs that monomethylate resveratrol to yield pinostilbene have been described so far. Here, we report the engineering of a resveratrol OMT from <i>Vitis vinifera</i> (VvROMT), which has the highest catalytic efficiency in di-methylating resveratrol to yield pterostilbene. In the absence of a crystal structure, we constructed a three-dimensional protein model of VvROMT and identified four critical binding site residues by applying different in silico approaches. We performed point mutations in these positions generating W20A, F24A, F311A, and F318A variants, which greatly reduced resveratrol’s enzymatic conversion. Then, we rationally designed eight variants through comparison of the binding site residues with other stilbene OMTs. We successfully modified the native substrate selectivity of VvROMT. Variant L117F/F311W showed the highest conversion to pinostilbene, and variant L117F presented an overall increase in enzymatic activity. Our results suggest that VvROMT has potential for the tailor-made production of stilbenes.https://www.mdpi.com/1422-0067/22/9/4345enzyme engineeringO-methyltransferasespinostilbeneprotein modelssubstrate selectivitystilbenes |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Daniela P. Herrera Andrea M. Chánique Ascensión Martínez-Márquez Roque Bru-Martínez Robert Kourist Loreto P. Parra Andreas Schüller |
spellingShingle |
Daniela P. Herrera Andrea M. Chánique Ascensión Martínez-Márquez Roque Bru-Martínez Robert Kourist Loreto P. Parra Andreas Schüller Rational Design of Resveratrol O-methyltransferase for the Production of Pinostilbene International Journal of Molecular Sciences enzyme engineering O-methyltransferases pinostilbene protein models substrate selectivity stilbenes |
author_facet |
Daniela P. Herrera Andrea M. Chánique Ascensión Martínez-Márquez Roque Bru-Martínez Robert Kourist Loreto P. Parra Andreas Schüller |
author_sort |
Daniela P. Herrera |
title |
Rational Design of Resveratrol O-methyltransferase for the Production of Pinostilbene |
title_short |
Rational Design of Resveratrol O-methyltransferase for the Production of Pinostilbene |
title_full |
Rational Design of Resveratrol O-methyltransferase for the Production of Pinostilbene |
title_fullStr |
Rational Design of Resveratrol O-methyltransferase for the Production of Pinostilbene |
title_full_unstemmed |
Rational Design of Resveratrol O-methyltransferase for the Production of Pinostilbene |
title_sort |
rational design of resveratrol o-methyltransferase for the production of pinostilbene |
publisher |
MDPI AG |
series |
International Journal of Molecular Sciences |
issn |
1661-6596 1422-0067 |
publishDate |
2021-04-01 |
description |
Pinostilbene is a monomethyl ether analog of the well-known nutraceutical resveratrol. Both compounds have health-promoting properties, but the latter undergoes rapid metabolization and has low bioavailability. O-methylation improves the stability and bioavailability of resveratrol. In plants, these reactions are performed by O-methyltransferases (OMTs). Few efficient OMTs that monomethylate resveratrol to yield pinostilbene have been described so far. Here, we report the engineering of a resveratrol OMT from <i>Vitis vinifera</i> (VvROMT), which has the highest catalytic efficiency in di-methylating resveratrol to yield pterostilbene. In the absence of a crystal structure, we constructed a three-dimensional protein model of VvROMT and identified four critical binding site residues by applying different in silico approaches. We performed point mutations in these positions generating W20A, F24A, F311A, and F318A variants, which greatly reduced resveratrol’s enzymatic conversion. Then, we rationally designed eight variants through comparison of the binding site residues with other stilbene OMTs. We successfully modified the native substrate selectivity of VvROMT. Variant L117F/F311W showed the highest conversion to pinostilbene, and variant L117F presented an overall increase in enzymatic activity. Our results suggest that VvROMT has potential for the tailor-made production of stilbenes. |
topic |
enzyme engineering O-methyltransferases pinostilbene protein models substrate selectivity stilbenes |
url |
https://www.mdpi.com/1422-0067/22/9/4345 |
work_keys_str_mv |
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1721515284827734016 |