scholarly journals Efficient Production of Enantiopure d-Lysine from l-Lysine by a Two-Enzyme Cascade System

Catalysts ◽  
2016 ◽  
Vol 6 (11) ◽  
pp. 168 ◽  
Author(s):  
Xin Wang ◽  
Li Yang ◽  
Weijia Cao ◽  
Hanxiao Ying ◽  
Kequan Chen ◽  
...  
Keyword(s):  
Efficient Production ◽  
Cascade System ◽  
Enzyme Cascade

scholarly journals A multi-enzyme cascade for efficient production of d-p-hydroxyphenylglycine from l-tyrosine

2021 ◽  
Vol 8 (1) ◽  
Author(s):  
Xu Tan ◽  
Sheng Zhang ◽  
Wei Song ◽  
Jia Liu ◽  
Cong Gao ◽  
...  
Keyword(s):  
Amino Acids ◽  
Specific Activity ◽  
Enantiomeric Excess ◽  
Efficient Production ◽  
Enzyme Cascade ◽  
Rate Limiting Step ◽  
Limiting Step ◽  
Rotation Strategy ◽  
Rate Limiting

AbstractIn this study, a four-enzyme cascade pathway was developed and reconstructed in vivo for the production of d-p-hydroxyphenylglycine (D-HPG), a valuable intermediate used to produce β-lactam antibiotics and in fine-chemical synthesis, from l-tyrosine. In this pathway, catalytic conversion of the intermediate 4-hydroxyphenylglyoxalate by meso-diaminopimelate dehydrogenase from Corynebacterium glutamicum (CgDAPDH) was identified as the rate-limiting step, followed by application of a mechanism-guided “conformation rotation” strategy to decrease the hydride-transfer distance d(C6HDAP−C4NNADP) and increase CgDAPDH activity. Introduction of the best variant generated by protein engineering (CgDAPDHBC621/D120S/W144S/I169P with 5.32 ± 0.85 U·mg−1 specific activity) into the designed pathway resulted in a D-HPG titer of 42.69 g/L from 50-g/L l-tyrosine in 24 h, with 92.5% conversion, 71.5% isolated yield, and > 99% enantiomeric excess in a 3-L fermenter. This four-enzyme cascade provides an efficient enzymatic approach for the industrial production of D-HPG from cheap amino acids.

Efficient One-Pot Synthesis of Cytidine 5′-Monophosphate Using an Extremophilic Enzyme Cascade System

2020 ◽  
Vol 68 (34) ◽  
pp. 9188-9194
Author(s):  
Zonglin Li ◽  
Xiao Ning ◽  
Yiran Zhao ◽  
Xiaodan Zhang ◽  
Chun Xiao ◽  
...  
Keyword(s):  
Cascade System ◽  
One Pot ◽  
One Pot Synthesis ◽  
Enzyme Cascade

A DNAzyme cascade for amplified detection of intracellular miRNA

2020 ◽  
Vol 56 (70) ◽  
pp. 10163-10166
Author(s):  
Yanan Wu ◽  
Jing Li ◽  
Ke Quan ◽  
Xiangxian Meng ◽  
Xiaohai Yang ◽  
...  
Keyword(s):  
Cascade Reaction ◽  
Cascade System ◽  
Enzyme Cascade ◽  
Natural Enzyme

Inspired by the natural enzyme cascade reaction, an artificial DNAzyme cascade system is developed for the amplified detection of intracellular miR-141.

scholarly journals Efficient Production of 3′-Sialyllactose by Single Whole-Cell in One-Pot Biosynthesis

Processes ◽  
2021 ◽  
Vol 9 (6) ◽  
pp. 932
Author(s):  
Zhongkui Li ◽  
Xiangsong Chen ◽  
Zhijian Ni ◽  
Lixia Yuan ◽  
Lijie Sun ◽  
...  
Keyword(s):  
Surfactant Concentration ◽  
Enzyme System ◽  
Efficient Production ◽  
Optimal Conditions ◽  
One Pot ◽  
Whole Cell ◽  
Economic Competitiveness ◽  
Enzyme Cascade ◽  
L System ◽  
Conversion Temperature

Sialyllactose (SL) is one of the most important acidic oligosaccharides in human milk, which plays an important role in the health of infants. In this work, an efficient multi-enzyme cascade was developed in a single whole cell to produce 3′-SL. We constructed two compatible plasmids with double cloning sites to co-express four genes. Different combinations were assessed to verify the optimal catalytic ability. Then, the conversion temperature, pH, and stability under the optimal temperature and pH were investigated. Moreover, the optimal conversion conditions and surfactant concentration were determined. By using the optimal conditions (35 °C, pH 7.0, 20 mM polyphosphate, 10 mM cytidine monophosphate (CMP), 20 mM MgCl2), 25 mL and 4 L conversion systems were carried out to produce 3′-SL. Similar results were obtained between different volume conversion reactions, which led the maximum production of 3′-SL to reach 53 mM from 54.2 mM of sialic acid (SA) in the 25 mL system and 52.8 mM of 3′-SL from 53.8 mM of SA in the 4 L system. These encouraging results demonstrate that the developed single whole-cell multi-enzyme system exhibits great potential and economic competitiveness for the manufacture of 3′-SL.

A two-enzyme cascade system for the bio-production of spermidine from putrescine

2021 ◽  
Vol 504 ◽  
pp. 111439
Author(s):  
Yi Liu ◽  
Xing Guo ◽  
Xin Wang ◽  
Kequan Chen ◽  
Pingkai Ouyang
Keyword(s):  
Cascade System ◽  
Enzyme Cascade

scholarly journals A Four-Step Enzymatic Cascade for Efficient Production of L-Phenylglycine from Biobased L-Phenylalanine

2022 ◽  
Author(s):  
Yuling Zhu ◽  
Jifeng Yuan
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Amino Acid ◽  
Industrial Applications ◽  
Efficient Production ◽  
E Coli ◽  
Amycolatopsis Orientalis ◽  
Enzyme Cascade ◽  
Pharmaceutical Industries ◽  
Rate Limiting

Enantiopure amino acids are of particular interest in the agrochemical and pharmaceutical industries. Here, we reported a multi-enzyme cascade for efficient production of L-phenylglycine (L-Phg) from biobased L-phenylalanine (L-Phe). We first attempted to engineer Escherichia coli for expressing L-amino acid deaminase (LAAD) from Proteus mirabilis, hydroxymandelate synthase (HmaS) from Amycolatopsis orientalis, (S)-mandelate dehydrogenase (SMDH) from Pseudomonas putida, the endogenous aminotransferase (AT) encoded by ilvE and L-glutamate dehydrogenase (GluDH) from E. coli. However, 10 mM L-Phe only afforded the synthesis of 7.21 mM L-Phg. The accumulation of benzoylformic acid suggested that the transamination step might be rate-limiting. We next used leucine dehydrogenase (LeuDH) from Bacillus cereus to bypass the use of L-glutamate as amine donor, and 40 mM L-Phe gave 39.97 mM (6.04 g/L) L-Phg, reaching 99.9% conversion. In summary, this work demonstrated a concise four-step enzymatic cascade for the L-Phg synthesis from biobased L-Phe, with a potential for future industrial applications.

scholarly journals Synthesis of Sitagliptin Intermediate by a Multi-Enzymatic Cascade System Using Lipase and Transaminase With Benzylamine as an Amino Donor

2021 ◽  
Vol 9 ◽  
Author(s):  
Taresh P. Khobragade ◽  
Sharad Sarak ◽  
Amol D. Pagar ◽  
Hyunwoo Jeon ◽  
Pritam Giri ◽  
...  
Keyword(s):  
Inhibitory Effect ◽  
Cell System ◽  
Buffer System ◽  
Cascade System ◽  
Whole Cell ◽  
Enzyme Cascade ◽  
Engineering Strategy ◽  
Amino Donor ◽  
Ph Buffer ◽  
Substrate Concentrations

Herein, we report the development of a multi-enzyme cascade using transaminase (TA), esterase, aldehyde reductase (AHR), and formate dehydrogenase (FDH), using benzylamine as an amino donor to synthesize the industrially important compound sitagliptin intermediate. A panel of 16 TAs was screened using ethyl 3-oxo-4-(2,4,5-trifluorophenyl) butanoate as a substrate (1). Amongst these enzymes, TA from Roseomonas deserti (TARO) was found to be the most suitable, showing the highest activity towards benzylamine (∼70%). The inhibitory effect of benzaldehyde was resolved by using AHR from Synechocystis sp. and FDH from Pseudomonas sp., which catalyzed the conversion of benzaldehyde to benzyl alcohol at the expense of NAD(P)H. Reaction parameters, such as pH, buffer system, and concentration of amino donor, were optimized. A single whole-cell system was developed for co-expressing TARO and esterase, and the promoter engineering strategy was adopted to control the expression level of each biocatalyst. The whole-cell reactions were performed with varying substrate concentrations (10–100 mM), resulting in excellent conversions (ranging from 72 to 91%) into the desired product. Finally, the applicability of this cascade was highlighted on Gram scale, indicating production of 70% of the sitagliptin intermediate with 61% isolated yield. The protocol reported herein may be considered an alternative to existing methods with respect to the use of cheaper amine donors as well as improved synthesis of (R) and (S) enantiomers with the use of non-chiral amino donors.

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