scholarly journals Protein engineering of Saccharomyces cerevisiae transporter Pdr5p identifies key residues that impact Fusarium mycotoxin export and resistance to inhibition

2016 ◽  
Vol 5 (6) ◽  
pp. 979-991 ◽  
Author(s):  
Amanda B. Gunter ◽  
Anne Hermans ◽  
Whynn Bosnich ◽  
Douglas A. Johnson ◽  
Linda J. Harris ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Protein Engineering ◽  
Fusarium Mycotoxin ◽  
Key Residues

scholarly journals Conversion of Waste Agriculture Biomass to Bioethanol by Recombinant Saccharomyces cerevisiae

2010 ◽  
Vol 2 (2) ◽  
pp. 351-361
Author(s):  
A. A. Saleh ◽  
S. Hamdan ◽  
N. Annaluru ◽  
S. Watanabe ◽  
M. R. Rahman ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Enzyme Activity ◽  
Protein Engineering ◽  
Agricultural Waste ◽  
Xylitol Dehydrogenase ◽  
Ethanol Conversion ◽  
Waste Biomass ◽  
Wild Type ◽  
Coenzyme Specificity ◽  
Recombinant Yeasts

Agricultural waste biomass has already been transferred to bioethanol and used as energy related products, although many issues such as efficiency and productivity still to be overcome. In this study, the protein engineering was applied to generate enzymes with completely reversed coenzyme specificity and developed recombinant yeasts containing those engineered enzymes for construction of an efficient biomass-ethanol conversion system. Recombinant yeasts were constructed with the genes encoding a wild type xylose reductase (XR) and the protein engineered xylitol dehydrogenase (XDH) (with NADP) of Pichia stipitis.  These recombinant yeasts were characterized based on the enzyme activity and fermentation ability of xylose to ethanol. The protein engineered enzymes were expressed significantly in Saccharomyces cerevisiae as judged by the enzyme activity in vitro. Ethanol fermentation was measured in batch culture under anaerobic conditions. The significant enhancement was found in Y-ARS strain, in which NADP+-dependent XDH was expressed; 85% decrease of unfavorable xylitol excretion with 26% increased ethanol production, when compared with the reference strain expressing the wild-type XDH.  Keywords: Agricultural waste biomass; Protein engineering; Xylitol dehydrogenase; Xylose-fermentation; Eethanol production. © 2010 JSR Publications. ISSN: 2070-0237 (Print); 2070-0245 (Online). All rights reserved. DOI: 10.3329/jsr.v2i2.2882               J. Sci. Res. 2 (2), 351-361 (2010) 

scholarly journals A deeper understanding of the spontaneous derepression of the URA3 gene in MaV203 Saccharomyces cerevisiae and its implications for protein engineering and the reverse two‐hybrid system

Yeast ◽  
2019 ◽  
Vol 36 (12) ◽  
pp. 701-710
Author(s):  
David Cortens ◽  
Rebekka Hansen ◽  
Geert‐Jan Graulus ◽  
Erik Steen Redeker ◽  
Peter Adriaensens ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Protein Engineering ◽  
Hybrid System ◽  
Ura3 Gene ◽  
Two Hybrid

Degradation of the minor Fusarium mycotoxin beauvericin by intracellular enzymes of Saccharomyces cerevisiae

Food Control ◽  
2013 ◽  
Vol 33 (2) ◽  
pp. 352-358 ◽  
Author(s):  
G. Meca ◽  
A. Ritieni ◽  
T. Zhou ◽  
X.Z. Li ◽  
J. Mañes
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Fusarium Mycotoxin ◽  
Intracellular Enzymes

scholarly journals Protein Engineering for Enhanced Enantioselectivity of Carboxylesterase Est924 Toward Ethyl 2-Arylpropionates

2021 ◽  
Author(s):  
Xiaolong Liu ◽  
Meng Zhao ◽  
Xinjiong Fan ◽  
Yao Fu
Keyword(s):  
Protein Engineering ◽  
Ethyl Ester ◽  
Rational Design ◽  
Kinetic Resolution ◽  
Structural Information ◽  
Careful Analysis ◽  
Chiral Drugs ◽  
Anti Inflammatory ◽  
Increasing Demand ◽  
Key Residues

Abstract BackgroundNon-steroidal anti-inflammatory drugs (NSAIDs) are the world's most used drugs with anti-inflammatory, analgesic and antipyretic effects. 2-arylpropionate drugs are a very important class of chiral drugs in NSAIDs. In the pharmaceutical industry, there is an increasing demand for esterase-mediated chiral resolution of racemic 2-arylpropionate esters due to the high activity and low side effects of the (S)-enantiomers. Esterases are important biocatalysts for chemical synthesis owing to their enantioselectivity, regioselectivity, broad substrate specificity and no need for cofactors. Several bHSL family esterases have been used to resolve racemic 2-arylpropionates. However, each reported enzyme was able to synthesize only one NSAIDs drug, which does not meet the industry requirement for catalysts and limits their application.ResultsThrough careful analysis of the structural information and molecular docking, we present the discovery of key residues that controlled the enantioselectivity of bHSL carboxylesterases to ethyl 2-arylpropionates. Est924 was identified as a bHSL family esterase and a promising catalyst for kinetic resolution of racemic ethyl 2-arylpropionates with sight (R)-stereopreference. Using Est924 as the starting enzyme, protein engineering at these key residues was conducted, and the substitution of A203 was shown to affect the enantioselectivity. The stereopreference of the mutant M1(A203W) was inverted to ethyl 2-(S)-arylpropionates, and this stereopreference was further improved in variant M3(I202F/A203W/G208F). In addition, the optimal variant, M3, was also suitable for the resolution of ibuprofen ethyl ester and ketoprofen ethyl ester, and their efficient (S)-isomers were synthesized.ConclusionsOur results suggested that Est924 variants could kinetically resolve economically important racemates for industrial production and further offer the opportunity for the rational design of enzyme enantioselectivity.

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.

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