Yingying Zhu,Dawei Ni,Zeyu Li,Zhebin Hao,Liang Wang,Wanmeng Mu
Indigo is an important blue pigment widely used in textile, food, and medicine industries. Biological production of indigo attracts increasing attention recently. Cell factory production of indigo encounters the problem of the toxicity of the precursor indole. Enzymatic production is the alternative biological approach, however, NADPH regeneration should be solved. In this study, flavin-containing monooxygenase from Methylophaga aminisulfidivorans was used for enzymatic production of indigo from indole and formate dehydrogenase from Pseudomonas sp. 101 was co-expressed for NADPH regeneration. Indigo production was enhanced by combination of molecular modification, promoter engineering, and translation initiation region engineering. Finally, 0.183 g/L of indigo was produced from 0.5 g/L of indole and 0.5 mM of sodium formate, with the conversion ratio of 32.5 %. This study demonstrates a feasible and effective strategy for enzymatic production of indigo.
Enzymatic preparation of rare sugars as an alternative to traditional sweeteners is an effective strategy to achieve a low-calorie healthy diet. Ribose-5-phosphate isomerase B (RpiB) is a key enzyme in the non-oxidative branch of the catalytic pentose phosphate pathway. Here, we investigated the potential of Curtobacterium flaccumfaciens ZXL1 ( C. flaccumfaciens ZXL1) derived RpiB (CfRpiB) in D-allose preparation. The optimal reaction conditions for recombinant CfRpiB were found experimentally to be pH 7.0, 55 °C, and no metal ions. The kinetic parameters K m , k cat , and catalytic efficiency k ca t / K m were 320 mM, 4769 s −1 , and 14.9 mM −1 s −1 respectively. The conversion of D-allulose by purified enzyme (1 g L −1 ) to D-allose was 13% within 1 h. In addition, homology modeling and molecular docking were used to predict the active site residues: Asp13, Asp14, Cys72, Gly73, Thr74, Gly77, Asn106, and Lys144.
Anlian Zhu,Wanlu Feng,Zhiyong Li,Shuang Cheng,Qianhan Chen,Dongshuang Fan,Yuanyang Guo,Lingjun Li,Jianji Wang
The great challenges of modern industry and the environment make it important to develop sustainable energy resources with low cost. In this work, a cleaner enzymatic procedure for biodiesel production was developed through the utilization of a biocompatible and hydrophilic ionic liquid [Choline][H2PO4]. This ionic liquid can be synthesized from cheap raw materials through simple neutralization procedures, and it has been proved to be well biocompatible. The utilization of this ionic liquid in Novozym 435 catalyzed biodiesel production makes the reaction and work-up procedures very simple, because its hydrophilicity can lead to the implementation of a pseudo homogeneous reaction and then heterogeneous separation. Various oil resources such as triolein, sunflower oil and castor oil can all be converted to biodiesels with high yields. After the completion of reaction, both the ionic liquid and Novozym 435 can be recycled and reutilized for at least five cycles without a significant activity decrease. In addition, this reaction system can be conveniently scaled up to the multi-gram level with high efficiency and feasible separation. Overall, the above mentioned benefits make this ionic liquid based enzymatic system cleaner for the production of biodiesel and promising for further industrial applications.
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