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Regioselective Hydroxylation of Stilbenes by Engineered Cytochrome P450 from Thermobifida fusca YX
Author(s) -
Rühlmann Ansgar,
Antovic Dragutin,
Müller Thomas J. J.,
Urlacher Vlada B.
Publication year - 2017
Publication title -
advanced synthesis and catalysis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.541
H-Index - 155
eISSN - 1615-4169
pISSN - 1615-4150
DOI - 10.1002/adsc.201601168
Subject(s) - chemistry , hydroxylation , monooxygenase , resveratrol , regioselectivity , cytochrome p450 , stereochemistry , enzyme , biocatalysis , biochemistry , catalysis , reaction mechanism
Since the past decades, the plant stilbenoid resveratrol has gained significant attention of the general public as well as the research community due to its versatile medicinal properties. Apart from resveratrol, there is also an increasing interest in other plant stilbenoids because of their different potential biological activities. In order to meet the increasing demand for stilbenoids, alternative and sustainable approaches for their production are needed. We identified the cytochrome P450 monooxygenase 154E1 from Thermobifida fusca YX (CYP154E1) which enables the synthesis of ( E )‐4,4′‐dihydroxystilbene via direct double hydroxylation of ( E )‐stilbene. The construction of a triple mutant led to a more than six‐fold increased catalytic efficiency compared to the wild type enzyme. CYP154E1 and variants thereof accepted not only ( E )‐stilbene but also possessed remarkable activity towards ortho ‐ and meta ‐substituted hydroxystilbenes leading to resveratrol, ( E )‐2,4′‐dihydroxystilbene, ( E )‐2,4′,5‐trihydroxystilbene and ( E )‐3,4′‐dihydroxystilbene. The combination of protein engineering and the use of methyl‐β‐cyclodextrin as substrate solubilizing agent resulted in product titers of up to 4.2 g L −1 and enzyme total turnover numbers (TTN) of up to 20,000.