P212A Mutant of Dihydrodaidzein Reductase Enhances (
            <i>S</i>
            )-Equol Production and Enantioselectivity in a Recombinant Escherichia coli Whole-Cell Reaction System

Lee, Pyung‐Gang; Kim, Joonwon; Kim, Eun-Jung; Jung, E.; Pandey, Bishnu Prasad; Kim, Byung‐Gee

doi:10.1128/aem.03584-15

Cited by 29 publications

(49 citation statements)

References 41 publications

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“…No equol production was detected under shaking at 200 rpm (data not shown) and this is consistent with previous reports ( Lee et al, 2016 ). Static culture conditions were utilized to detect the equol production in this study.…”

Section: Resultssupporting

confidence: 93%

“…The gene product of L- dznr is responsible for converting daidzein into ( R )-dihydrodaidzein; the L- ddrc gene product converts ( R )-dihydrodaidzein into ( S )-dihydrodaidzein; the L- dhdr gene product converts ( S )-dihydrodaidzein into trans -tetrahydrodaidze; and the L- thdr gene product converts trans -tetrahydrodaidzein into ( S )-equol ( Shimada et al, 2011 , 2012 ). Lee et al (2016) constructed a recombinant Escherichia coli BL21 strain which can produce (S)-equol in vitro . However, Vázquez et al (2017) reported that isoflavone-derived compounds like ( S )-equol have the ability to inhibit the growth from many bacteria species.…”

Section: Introductionmentioning

confidence: 99%

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To Construct an Engineered (S)-Equol Resistant E. coli for in Vitro (S)-Equol Production

et al. 2018

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(S)-equol is one of the major metabolites of daidzein that is produced by human and animal gut bacteria. Most of the physiological functions of soybean isoflavones, such as anti-oxidative activity, anti-cancer activity, and cardiovascular protection have been ascribed to (S)-equol. However, only 30–50% people contain this kind of equol-producing bacteria, and therefore are able to convert daidzein to (S)-equol. Administration of (S)-equol may be more beneficial than soybean isoflavones. The aim of this study was to construct an engineered (S)-equol resistant Escherichia coli to enhance (S)-equol production in vitro. First, transposon mutagenesis libraries were constructed and screened to isolate the (S)-equol resistant mutant E. coli strain BL21 (ydiS) in order to overcome the inhibitory effects of (S)-equol on bacterial growth. Bacterial full genome scan sequencing and in vitro overexpression results revealed that the ydiS gene was responsible for this resistance. Second, the (S)-equol-producing genes L-dznr, L-ddrc, L-dhdr, and L-thdr of Lactococcus strain 20–92 were synthesized and cloned into compatible vectors, pETDuet-1 and pCDFDuet-1. These plasmids were subsequently transformed into BL21 (DE3) and its mutant BL21 (ydiS). Both engineered BL21 (DE3) and BL21 (ydiS) could use daidzein as substrate to produce (S)-equol under both anaerobic and aerobic conditions. As expected, engineered BL21 (ydiS) had faster growth rates than BL21 (DE3) when supplemented with high concentrations of (S)-equol. The yield and the daidzein utilization ratio were higher for engineered BL21 (ydiS). Interestingly, engineered BL21 (ydiS) was able to convert daidzein to (S)-equol efficiently under aerobic conditions, providing a convenient method for (S)-equol production in vitro. In addition, a two-step method was developed to produce (S)-equol using daidzin as substrate.

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Section: Resultssupporting

confidence: 93%

Section: Introductionmentioning

confidence: 99%

To Construct an Engineered (S)-Equol Resistant E. coli for in Vitro (S)-Equol Production

et al. 2018

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“…24 ). Different anaerobic bacteria can reduce daidzein to S -equol through three different reductase enzymes and one dihydrodaidzein racemase ( Shimada et al, 2011 ; Lee et al, 2016 ). Genetic polymorphism in equol production has been demonstrated, which in part may explain the variability in relieving menopausal symptoms ( Hong et al, 2012 ).…”

Section: Botanicals Used By Menopausal Womenmentioning

confidence: 99%

Botanicals and Their Bioactive Phytochemicals for Women’s Health

et al. 2016

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Botanical dietary supplements are increasingly popular for women’s health, particularly for older women. The specific botanicals women take vary as a function of age. Younger women will use botanicals for urinary tract infections, especially Vaccinium macrocarpon (cranberry), where there is evidence for efficacy. Botanical dietary supplements for premenstrual syndrome (PMS) are less commonly used, and rigorous clinical trials have not been done. Some examples include Vitex agnus-castus (chasteberry), Angelica sinensis (dong quai), Viburnum opulus/prunifolium (cramp bark and black haw), and Zingiber officinale (ginger). Pregnant women have also used ginger for relief from nausea. Natural galactagogues for lactating women include Trigonella foenum-graecum (fenugreek) and Silybum marianum (milk thistle); however, rigorous safety and efficacy studies are lacking. Older women suffering menopausal symptoms are increasingly likely to use botanicals, especially since the Women’s Health Initiative showed an increased risk for breast cancer associated with traditional hormone therapy. Serotonergic mechanisms similar to antidepressants have been proposed for Actaea/Cimicifuga racemosa (black cohosh) and Valeriana officinalis (valerian). Plant extracts with estrogenic activities for menopausal symptom relief include Glycine max (soy), Trifolium pratense (red clover), Pueraria lobata (kudzu), Humulus lupulus (hops), Glycyrrhiza species (licorice), Rheum rhaponticum (rhubarb), Vitex agnus-castus (chasteberry), Linum usitatissimum (flaxseed), Epimedium species (herba Epimedii, horny goat weed), and Medicago sativa (alfalfa). Some of the estrogenic botanicals have also been shown to have protective effects against osteoporosis. Several of these botanicals could have additional breast cancer preventive effects linked to hormonal, chemical, inflammatory, and/or epigenetic pathways. Finally, although botanicals are perceived as natural safe remedies, it is important for women and their healthcare providers to realize that they have not been rigorously tested for potential toxic effects and/or drug/botanical interactions. Understanding the mechanism of action of these supplements used for women’s health will ultimately lead to standardized botanical products with higher efficacy, safety, and chemopreventive properties.

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“…DHDR was thought to be the rate‐determining enzyme in the recombinant strain biotransformation process with low daidzein concentrations (below 1mM). Using site‐directed mutation of DHDR with P212A, Lee et al () improved the ( S )‐equol productivity of the recombinant strain. As observed, THDR was capable merely to convert (3 S ,4 R )‐ trans ‐tetrahydrodaidzein to ( S )‐equol involving a radical intermediate.…”

Section: Approaches For Equol Productionmentioning

confidence: 99%

“…Lee et al established the heterologous co‐expression of the four‐step enzyme process from Slackia isoflavoniconvertens in a recombinant E.coli . Subsequent to that, they improved the ( S )‐equol yield to 69.8 mg L −1 hr −1 using the site directed mutation of DHDR with P212A in the recombinant E.coli (Lee et al, ). Ultimately, the yield reached 1.22 g/L of ( S )‐equol from 1.27 g/L of daidzein, by adding 5% (w/v) PVP‐40k and 5% (v/v) DMSO as co‐solvents in the whole‐cell biotransformation system (Lee et al, ; Figure ).…”

Section: Approaches For Equol Productionmentioning

confidence: 99%

Advances in exploring equol production and application

2019

J Food Process Preserv

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The soybean isoflavone daidzein has been extensively studied. Equol represents the main active metabolite of daidzein in vivo. Due to its broadly biological activities, the preparation and characterization of equol have attracted a great deal of interest. In this review, the focus of attention is placed on the approaches to equol production, including the total synthesis of equol, equol from natural bacteria fermentation and synthetic biology for equol production. It is expected that the industrial production of equol with high yield could be realized in an economic way in the future. In addition, pharmacological activities and clinical applications of equol are also discussed to gain a systematically understanding as to the prospect of development for equol. Practical applications Equol is the main active metabolite of daidzein in vivo and considered to be the most active metabolite of the soybean isoflavones. It has broadly biological activities, including antitumor, antioxidative properties, cardiovascular protection, and relieve menopausal symptoms. However, only 25%–30% of young people can excrete equol in vivo when supplied with soy foods. So, it is necessary to develop efficient methods for equol preparation. Meanwhile, more systematic researches for the pharmacological activity of equol are also required. As mentioned above, this manuscript entitled “Advances in exploring Equol production and application,” hoping that this review should draw widespread attention.

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P212A Mutant of Dihydrodaidzein Reductase Enhances ( S )-Equol Production and Enantioselectivity in a Recombinant Escherichia coli Whole-Cell Reaction System

Cited by 29 publications

References 41 publications

To Construct an Engineered (S)-Equol Resistant E. coli for in Vitro (S)-Equol Production

To Construct an Engineered (S)-Equol Resistant E. coli for in Vitro (S)-Equol Production

Botanicals and Their Bioactive Phytochemicals for Women’s Health

Advances in exploring equol production and application

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