scholarly journals Engineering of Yeast Old Yellow Enzyme OYE3 Enables Its Capability Discriminating of (E)-Citral and (Z)-Citral

Molecules ◽  
2021 ◽  
Vol 26 (16) ◽  
pp. 5040
Author(s):  
Tairan Wang ◽  
Ran Wei ◽  
Yingting Feng ◽  
Lijun Jin ◽  
Yunpeng Jia ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Rational Design ◽  
Asymmetric Reduction ◽  
Glucose Dehydrogenase ◽  
Direct Synthesis ◽  
Cascade Reaction ◽  
Old Yellow Enzyme ◽  
Complete Reduction ◽  
Novel Approach ◽  
Double Substitution

The importance of yeast old yellow enzymes is increasingly recognized for direct asymmetric reduction of (E/Z)-citral to (R)-citronellal. As one of the most performing old yellow enzymes, the enzyme OYE3 from Saccharomyces cerevisiae S288C exhibited complementary enantioselectivity for the reduction of (E)-citral and (Z)-citral, resulting in lower e.e. value of (R)-citronellal in the reduction of (E/Z)-citral. To develop a novel approach for the direct synthesis of enantio-pure (R)-citronellal from the reduction of (E/Z)-citral, the enzyme OYE3 was firstly modified by semi-rational design to improve its (R)-enantioselectivity. The OYE3 variants W116A and S296F showed strict (R)-enantioselectivity in the reduction of (E)-citral, and significantly reversed the (S)-enantioselectivity in the reduction of (Z)-citral. Next, the double substitution of OYE3 led to the unique variant S296F/W116G, which exhibited strict (R)-enantioselectivity in the reduction of (E)-citral and (E/Z)-citral, but was not active on (Z)-citral. Relying on its capability discriminating (E)-citral and (Z)-citral, a new cascade reaction catalyzed by the OYE3 variant S296F/W116G and glucose dehydrogenase was developed, providing the enantio-pure (R)-citronellal and the retained (Z)-citral after complete reduction of (E)-citral.

scholarly journals Production of a Doubly Chiral Compound, (4R,6R)-4-Hydroxy-2,2,6-Trimethylcyclohexanone, by Two-Step Enzymatic Asymmetric Reduction

2003 ◽  
Vol 69 (2) ◽  
pp. 933-937 ◽  
Author(s):  
Masaru Wada ◽  
Ayumi Yoshizumi ◽  
Yumiko Noda ◽  
Michihiko Kataoka ◽  
Sakayu Shimizu ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Saccharomyces Cerevisiae ◽  
Double Bond ◽  
Enzymatic Synthesis ◽  
Asymmetric Reduction ◽  
Glucose Dehydrogenase ◽  
Enantiomeric Excess ◽  
Old Yellow Enzyme ◽  
Chiral Compound ◽  
E Coli

ABSTRACT A practical enzymatic synthesis of a doubly chiral key compound, (4R,6R)-4-hydroxy-2,2,6-trimethylcyclohexanone, starting from the readily available 2,6,6-trimethyl-2-cyclohexen-1,4-dione is described. Chirality is first introduced at the C-6 position by a stereoselective enzymatic hydrogenation of the double bond using old yellow enzyme 2 of Saccharomyces cerevisiae, expressed in Escherichia coli, as a biocatalyst. Thereafter, the carbonyl group at the C-4 position is reduced selectively and stereospecifically by levodione reductase of Corynebacterium aquaticum M-13, expressed in E. coli, to the corresponding alcohol. Commercially available glucose dehydrogenase was also used for cofactor regeneration in both steps. Using this two-step enzymatic asymmetric reduction system, 9.5 mg of (4R,6R)-4-hydroxy-2,2,6-trimethylcyclohexanone/ml was produced almost stoichiometrically, with 94% enantiomeric excess in the presence of glucose, NAD+, and glucose dehydrogenase. To our knowledge, this is the first report of the application of S. cerevisiae old yellow enzyme for the production of a useful compound.

scholarly journals Engineering the Enantioselectivity of Yeast Old Yellow Enzyme OYE2y in Asymmetric Reduction of (E/Z)-Citral to (R)-Citronellal

Molecules ◽  
2019 ◽  
Vol 24 (6) ◽  
pp. 1057 ◽  
Author(s):  
Xiangxian Ying ◽  
Shihua Yu ◽  
Meijuan Huang ◽  
Ran Wei ◽  
Shumin Meng ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Protein Engineering ◽  
Active Site ◽  
Active Sites ◽  
Asymmetric Reduction ◽  
Saturation Mutagenesis ◽  
Old Yellow Enzyme ◽  
Docking Analysis ◽  
Site Saturation ◽  
Low Activity

The members of the Old Yellow Enzyme (OYE) family are capable of catalyzing the asymmetric reduction of (E/Z)-citral to (R)-citronellal—a key intermediate in the synthesis of L-menthol. The applications of OYE-mediated biotransformation are usually hampered by its insufficient enantioselectivity and low activity. Here, the (R)-enantioselectivity of Old Yellow Enzyme from Saccharomyces cerevisiae CICC1060 (OYE2y) was enhanced through protein engineering. The single mutations of OYE2y revealed that the sites R330 and P76 could act as the enantioselectivity switch of OYE2y. Site-saturation mutagenesis was conducted to generate all possible replacements for the sites R330 and P76, yielding 17 and five variants with improved (R)-enantioselectivity in the (E/Z)-citral reduction, respectively. Among them, the variants R330H and P76C partly reversed the neral derived enantioselectivity from 32.66% e.e. (S) to 71.92% e.e. (R) and 37.50% e.e. (R), respectively. The docking analysis of OYE2y and its variants revealed that the substitutions R330H and P76C enabled neral to bind with a flipped orientation in the active site and thus reverse the enantioselectivity. Remarkably, the double substitutions of R330H/P76M, P76G/R330H, or P76S/R330H further improved (R)-enantioselectivity to >99% e.e. in the reduction of (E)-citral or (E/Z)-citral. The results demonstrated that it was feasible to alter the enantioselectivity of OYEs through engineering key residue distant from active sites, e.g., R330 in OYE2y.

scholarly journals Cascading Old Yellow Enzyme, Alcohol Dehydrogenase and Glucose Dehydrogenase for Selective Reduction of (E/Z)-Citral to (S)-Citronellol

Catalysts ◽  
2021 ◽  
Vol 11 (8) ◽  
pp. 931
Author(s):  
Yunpeng Jia ◽  
Qizhou Wang ◽  
Jingjing Qiao ◽  
Binbin Feng ◽  
Xueting Zhou ◽  
...  
Keyword(s):  
Alcohol Dehydrogenase ◽  
Glucose Dehydrogenase ◽  
Selective Reduction ◽  
Coli Strain ◽  
Severe Reaction ◽  
High Catalytic Activity ◽  
Old Yellow Enzyme ◽  
One Pot ◽  
Nadph Regeneration ◽  
By Products

Citronellol is a kind of unsaturated alcohol with rose-like smell and its (S)-enantiomer serves as an important intermediate for organic synthesis of (-)-cis-rose oxide. Chemical methods are commonly used for the synthesis of citronellol and its (S)-enantiomer, which suffers from severe reaction conditions and poor selectivity. Here, the first one-pot double reduction of (E/Z)-citral to (S)-citronellol was achieved in a multi-enzymatic cascade system: N-ethylmaleimide reductase from Providencia stuartii (NemR-PS) was selected to catalyze the selective reduction of (E/Z)-citral to (S)-citronellal, alcohol dehydrogenase from Yokenella sp. WZY002 (YsADH) performed the further reduction of (S)-citronellal to (S)-citronellol, meanwhile a variant of glucose dehydrogenase from Bacillus megaterium (BmGDHM6), together with glucose, drove efficient NADPH regeneration. The Escherichia coli strain co-expressing NemR-PS, YsADH, and BmGDHM6 was successfully constructed and used as the whole-cell catalyst. Various factors were investigated for achieving high conversion and reducing the accumulation of the intermediate (S)-citronellal and by-products. 0.4 mM NADP+ was essential for maintaining high catalytic activity, while the feeding of the cells expressing BmGDHM6 effectively eliminated the intermediate and by-products and shortened the reaction time. Under optimized conditions, the bio-transformation of 400 mM citral caused nearly complete conversion (>99.5%) to enantio-pure (S)-citronellol within 36 h, demonstrating promise for industrial application.

scholarly journals Construction of fast xylose-fermenting yeast based on industrial ethanol-producing diploid Saccharomyces cerevisiae by rational design and adaptive evolution

2013 ◽  
Vol 13 (1) ◽  
pp. 110 ◽  
Author(s):  
Liuyang Diao ◽  
Yingmiao Liu ◽  
Fenghui Qian ◽  
Junjie Yang ◽  
Yu Jiang ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Adaptive Evolution ◽  
Rational Design

Green asymmetric reduction of acetophenone derivatives: Saccharomyces cerevisiae and aqueous natural deep eutectic solvent

2018 ◽  
Vol 41 (2) ◽  
pp. 253-262 ◽  
Author(s):  
Manuela Panić ◽  
Doris Delač ◽  
Marin Roje ◽  
Ivana Radojčić Redovniković ◽  
Marina Cvjetko Bubalo
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Asymmetric Reduction ◽  
Deep Eutectic Solvent ◽  
Acetophenone Derivatives ◽  
Natural Deep Eutectic Solvent

scholarly journals Direct synthesis of aryl-annulated [c]carbazoles by gold(i)-catalysed cascade reaction of azide-diynes and arenes

2018 ◽  
Vol 9 (44) ◽  
pp. 8416-8425 ◽  
Author(s):  
Yuiki Kawada ◽  
Shunsuke Ohmura ◽  
Misaki Kobayashi ◽  
Wataru Nojo ◽  
Masaki Kondo ◽  
...  
Keyword(s):  
Direct Synthesis ◽  
Cascade Reaction ◽  
Azido Group

The gold-catalysed annulation of conjugated alkynes bearing an azido group with arenes gave annulated [c]carbazoles.

scholarly journals CRISPR-PCDup: a novel approach for simultaneous segmental chromosomal duplication in Saccharomyces cerevisiae

AMB Express ◽  
2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Naim Hassan ◽  
Yu Sasano ◽  
Shunta Kimura ◽  
Farhana Easmin ◽  
Keisuke Ekino ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Novel Approach ◽  
Chromosomal Duplication

scholarly journals Novel Approach in the Construction of Bioethanol-Producing Saccharomyces cerevisiae Hybrids

2019 ◽  
Vol 57 (1) ◽  
pp. 5-16 ◽  
Author(s):  
Anamarija Štafa ◽  
Andrea Pranklin ◽  
Ivan Krešimir Svetec ◽  
Božidar Šantek ◽  
Marina Svetec Miklenić ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Gene Targeting ◽  
Co2 Production ◽  
Production Test ◽  
Fermentation Inhibitors ◽  
Lignocellulosic Hydrolysates ◽  
Novel Approach ◽  
Ethanol Producer ◽  
High Ethanol ◽  
Selection Of

Bioethanol production from lignocellulosic hydrolysates requires a producer strain that tolerates both the presence of growth and fermentation inhibitors and high ethanol concentrations. Therefore, we constructed heterozygous intraspecies hybrid diploids of Saccharomyces cerevisiae by crossing two natural S. cerevisiae isolates, YIIc17_E5 and UWOPS87-2421, a good ethanol producer found in wine and a strain from the flower of the cactus Opuntia megacantha resistant to inhibitors found in lignocellulosic hydrolysates, respectively. Hybrids grew faster than parental strains in the absence and in the presence of acetic and levulinic acids and 2-furaldehyde, inhibitors frequently found in lignocellulosic hydrolysates, and the overexpression of YAP1 gene increased their survival. Furthermore, although originating from the same parental strains, hybrids displayed different fermentative potential in a CO2 production test, suggesting genetic variability that could be used for further selection of desirable traits. Therefore, our results suggest that the construction of intraspecies hybrids coupled with the use of genetic engineering techniques is a promising approach for improvement or development of new biotechnologically relevant strains of S. cerevisiae. Moreover, it was found that the success of gene targeting (gene targeting fidelity) in natural S. cerevisiae isolates (YIIc17_E5α and UWOPS87-2421α) was strikingly lower than in laboratory strains and the most frequent off-targeting event was targeted chromosome duplication.

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