Round, round we go – strategies for enzymatic cofactor regeneration

2020 ◽  
Vol 37 (10) ◽  
pp. 1316-1333 ◽  
Author(s):  
Silja Mordhorst ◽  
Jennifer N. Andexer
Keyword(s):  
Cofactor Regeneration ◽  
Regeneration System ◽  
Complex Molecules

Combined with an efficient and flexible regeneration system, cofactor-dependent enzymes can be used to selectively introduce modifications in complex molecules.

Cell-Free Total Biosynthesis of Plant Terpene Natural Products Using an Orthogonal Cofactor Regeneration System

ACS Catalysis ◽  
2021 ◽  
pp. 9898-9903
Author(s):  
Undramaa Bat-Erdene ◽  
John M. Billingsley ◽  
William C. Turner ◽  
Benjamin R. Lichman ◽  
Francesca M. Ippoliti ◽  
...  
Keyword(s):  
Natural Products ◽  
Cofactor Regeneration ◽  
Regeneration System

scholarly journals Construction and characterization of a novel glucose dehydrogenase-leucine dehydrogenase fusion enzyme for the biosynthesis of l-tert-leucine

2021 ◽  
Vol 20 (1) ◽  
Author(s):  
Langxing Liao ◽  
Yonghui Zhang ◽  
Yali Wang ◽  
Yousi Fu ◽  
Aihui Zhang ◽  
...  
Keyword(s):  
Glucose Dehydrogenase ◽  
Catalytic Efficiency ◽  
Space Time ◽  
Cofactor Regeneration ◽  
Enzyme Complex ◽  
Regeneration System ◽  
Regeneration Rate ◽  
Leucine Dehydrogenase ◽  
Fusion Enzyme ◽  
Space Time Yield

Abstract Background Biosynthesis of l-tert-leucine (l-tle), a significant pharmaceutical intermediate, by a cofactor regeneration system friendly and efficiently is a worthful goal all the time. The cofactor regeneration system of leucine dehydrogenase (LeuDH) and glucose dehydrogenase (GDH) has showed great coupling catalytic efficiency in the synthesis of l-tle, however the multi-enzyme complex of GDH and LeuDH has never been constructed successfully. Results In this work, a novel fusion enzyme (GDH–R3–LeuDH) for the efficient biosynthesis of l-tle was constructed by the fusion of LeuDH and GDH mediated with a rigid peptide linker. Compared with the free enzymes, both the environmental tolerance and thermal stability of GDH–R3–LeuDH had a great improved since the fusion structure. The fusion structure also accelerated the cofactor regeneration rate and maintained the enzyme activity, so the productivity and yield of l-tle by GDH–R3–LeuDH was all enhanced by twofold. Finally, the space–time yield of l-tle catalyzing by GDH–R3–LeuDH whole cells could achieve 2136 g/L/day in a 200 mL scale system under the optimal catalysis conditions (pH 9.0, 30 °C, 0.4 mM of NAD+ and 500 mM of a substrate including trimethylpyruvic acid and glucose). Conclusions It is the first report about the fusion of GDH and LeuDH as the multi-enzyme complex to synthesize l-tle and reach the highest space–time yield up to now. These results demonstrated the great potential of the GDH–R3–LeuDH fusion enzyme for the efficient biosynthesis of l-tle.

scholarly journals Demethylation of vanillic acid by recombinant LigM in a one-pot cofactor regeneration system

2016 ◽  
Vol 6 (21) ◽  
pp. 7729-7737 ◽  
Author(s):  
Elena Rosini ◽  
Paola D'Arrigo ◽  
Loredano Pollegioni
Keyword(s):  
Vanillic Acid ◽  
Cofactor Regeneration ◽  
Methionine Synthase ◽  
Regeneration System ◽  
One Pot ◽  
Cofactor Recycling ◽  
System P

Recombinant LigM from Sphingobium SYK-6 and plant methionine synthase MetE enzyme efficiently convert vanillic acid into PCA with cofactor recycling.

scholarly journals Intensifying Electron Utilization by Surface-Anchored Rh Complex for Enhanced Nicotinamide Cofactor Regeneration and Photoenzymatic CO2 Reduction

Research ◽  
2021 ◽  
Vol 2021 ◽  
pp. 1-11
Author(s):  
Yuqing Cheng ◽  
Jiafu Shi ◽  
Yizhou Wu ◽  
Xueying Wang ◽  
Yiying Sun ◽  
...  
Keyword(s):  
Nicotinamide Adenine Dinucleotide ◽  
Cofactor Regeneration ◽  
Utilization Efficiency ◽  
Nicotinamide Adenine ◽  
Homogeneous Catalyst ◽  
Enzymatic Reactions ◽  
Regeneration System ◽  
Nadh Regeneration ◽  
Regeneration Efficiency ◽  
Reduced Nicotinamide Adenine Dinucleotide

Solar-driven photocatalytic regeneration of cofactors, including reduced nicotinamide adenine dinucleotide (NADH), reduced nicotinamide adenine dinucleotide phosphate (NADPH), and reduced flavin adenine dinucleotide (FADH2), could ensure the sustainable energy supply of enzymatic reactions catalyzed by oxidoreductases for the efficient synthesis of chemicals. However, the elevation of cofactor regeneration efficiency is severely hindered by the inefficient utilization of electrons transferred on the surface of photocatalysts. Inspired by the phenomenon of ferredoxin-NADP+ reductase (FNR) anchoring on thylakoid membrane, herein, a homogeneous catalyst of rhodium (Rh) complex, [Cp∗Rh(bpy)H2O]2+, was anchored on polymeric carbon nitride (PCN) mediated by a tannic acid/polyethyleneimine (TA/PEI) adhesive layer, acquiring PCN@TA/PEI-Rh core@shell photocatalyst. Illuminated by visible light, electrons were excited from the PCN core, then transferred through the TA/PEI shell, and finally captured by the surface-anchored Rh for instant utilization during the regeneration of NADH. The TA/PEI-Rh shell could facilitate the electron transfer from the PCN core and, more importantly, achieved ~1.3-fold elevation of electron utilization efficiency compared with PCN. Accordingly, the PCN@TA/PEI-Rh afforded the NADH regeneration efficiency of 37.8% after 20 min reaction under LED light (405 nm) illumination, over 1.5 times higher than PCN with free Rh. Coupling of the NADH regeneration system with formate dehydrogenase achieved continuous production of formate from carbon dioxide (CO2). Our study may provide a generic and effective strategy to elevate the catalytic efficiency of a photocatalyst through intensifying the electron utilization.

Efficient asymmetric synthesis of chiral alcohols using high 2-propanol tolerance alcohol dehydrogenase SmADH2 via an environmentally friendly TBCR system

2020 ◽  
Vol 10 (1) ◽  
pp. 70-78 ◽  
Author(s):  
Zeyu Yang ◽  
Hengwei Fu ◽  
Wenjie Ye ◽  
Youyu Xie ◽  
Qinghai Liu ◽  
...  
Keyword(s):  
Alcohol Dehydrogenase ◽  
Asymmetric Synthesis ◽  
Environmentally Friendly ◽  
Cofactor Regeneration ◽  
Chiral Alcohols ◽  
Regeneration System ◽  
High Substrate ◽  
System A ◽  
System P

Based on substrate-coupled cofactor regeneration system, a high 2-propanol tolerance SmADH2 together with TBCR system can synthesise structurally diverse chiral alcohols at a high substrate loading with only 1.25 equivalents of 2-propanol.

scholarly journals Construction and characterization of a novel glucose dehydrogenase-leucine dehydrogenase fusion enzyme for the biosynthesis of L-tert-leucine

2020 ◽  
Author(s):  
Langxing Liao ◽  
Yonghui Zhang ◽  
Yali Wang ◽  
Yousi Fu ◽  
Aihui Zhang ◽  
...  
Keyword(s):  
Glucose Dehydrogenase ◽  
Catalytic Efficiency ◽  
Space Time ◽  
Cofactor Regeneration ◽  
Enzyme Complex ◽  
Regeneration System ◽  
Regeneration Rate ◽  
Leucine Dehydrogenase ◽  
Fusion Enzyme ◽  
Space Time Yield

Abstract Background: Biosynthesis of L-tert-leucine (L-tle), a significant pharmaceutical intermediate, by a cofactor regeneration system friendly and efficiently is a worthful goal all the time. The cofactor regeneration system of leucine dehydrogenase (LeuDH) and glucose dehydrogenase (GDH) has showed great coupling catalytic efficiency in the synthesis of L-tle, however the multi-enzyme complex of GDH and LeuDH has never been constructed successfully.Results: In this work, a novel fusion enzyme (GDH-R3-LeuDH) for the efficient biosynthesis of L-tle was constructed by the fusion of LeuDH and GDH mediated with a rigid peptide linker. Compared with the free enzymes, both the environmental tolerance and thermal stability of GDH-R3-LeuDH had a great improved since the fusion structure. The fusion structure also accelerated the cofactor regeneration rate and maintained the enzyme activity, so the productivity and yeild of L-tle by GDH-R3-LeuDH was all enhanced by 2-fold. Finally, the space-time yield of L-tle catalyzing by GDH-R3-LeuDH whole cells could achieve 2136 g/L/d in a 200 mL scale system under the optimal catalysis conditions (pH 9.0, 30 °C, 0.4 mM of NAD+ and 500 mM of a substrate including trimethylpyruvic acid and glucose).Conclusions: It is the first report about the fusion of GDH and LeuDH as the multi-enzyme complex to synthesize L-tle and reach the highest space-time yield up to now. These results demonstrated the great potential of the GDH-R3-LeuDH fusion enzyme for the efficient biosynthesis of L-tle.

scholarly journals Developing an Ethanol Utilization Pathway based NADH Regeneration System in Escherichia coli

2021 ◽  
Vol 2 (9) ◽  
pp. 01-11
Author(s):  
Wenfa Ng
Keyword(s):  
Escherichia Coli ◽  
Glucose Dehydrogenase ◽  
Cofactor Regeneration ◽  
Regeneration System ◽  
Nadh Regeneration ◽  
Operating Modes ◽  
Theoretical Yield ◽  
Growing Cell ◽  
Biocatalytic Reaction ◽  
Utilization Pathway

Interests remain in searching for cofactor regeneration system with higher efficiency at lower substrate cost. Glucose dehydrogenase (GDH) system has been dominant in NADH regeneration, but it only has a theoretical yield of one NADH per glucose molecule. This work sought to explore the utility of a two-step ethanol utilization pathway (EUP) in pathway-based NADH regeneration. The pathway runs from ethanol to acetaldehyde and to acetyl-CoA with each step generating one NADH, that together results in a higher theoretical yield of two NADH per ethanol molecule. In this project, anaerobic biotransformation of ketone (acetophenone or butanone) to alcohol by cpsADH from Candida parapsilosis was used as readout for evaluating relative efficacy and operating modes for EUP cofactor regeneration in Escherichia coli BL21 (DE3). Experiment tests validated that EUP was more efficient than GDH in NADH regeneration. Further, growing cell delivered higher biotransformation efficiency compared to resting cell due to the driving force generated by cell growth. Finally, preculture or cultivation in M9 + 10 g/L ethanol medium delivered higher biotransformation efficiency compared to LB medium. Overall, EUP could help regenerate NADH in support of a biocatalytic reaction, and is more efficient in cofactor regeneration than GDH.

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