Multienzyme processes represent an important area of biocatalysis. Their efficiency can be enhanced by optimization of the stoichiometry of the biocatalysts. Here we present a workflow for maximizing the efficiency of a three‐enzyme system catalyzing a five‐step chemical conversion. Kinetic models of pathways with wild‐type or engineered enzymes were built, and the enzyme stoichiometry of each pathway was optimized. Mathematical modeling and one‐pot multienzyme experiments provided detailed insights into pathway dynamics, enabled the selection of a suitable engineered enzyme, and afforded high efficiency while minimizing biocatalyst loadings. Optimizing the stoichiometry in a pathway with an engineered enzyme reduced the total biocatalyst load by an impressive 56 %. Our new workflow represents a broadly applicable strategy for optimizing multienzyme processes. 相似文献
Enzyme cascade reactions play significant roles in bioelectrochemical processes because they permit more complex reactions. Co-immobilization of multienzyme on the electrode could help to facilitate substrate/intermediate transfer among different enzymes and electron transfer from enzyme active sites to the electrode with high stability and retrievability. Different co-immobilization strategies to construct multienzyme bioelectrodes have been widely reported, however, up to now, they have barely been reviewed. In this review, we focus on recent state-of-the-art techniques for constructing co-immobilized multienzyme electrodes including random and positional co-immobilization. Particular attention is given to strategies such as multienzyme complex and surface display. Cofactor co-immobilization on the electrode is also crucial for the enhancement of catalytic reaction and electron transfer, yet, few studies have been reported. The up-to-date advances in bioelectrochemical applications of multienzyme bioelectrodes are also presented. Finally, key challenges and future perspectives are discussed. 相似文献
Here we present the applicability of different imine reductases (IREDs) in micro‐aqueous reaction systems. Subjects of the study were the IREDs from Streptomyces aurantiacus (SaIR), Streptomyces sp. GF3587 (RGF3587IR), Streptomyces kanamyceticus (SkIR), Streptomyces ipomoeae 91‐03 (SiIR), Streptomyces sp. GF3546 (SGF3546IR), and Paenibacillus elgii B69 (PeIR). The IREDs were overexpressed in Escherichia coli (E. coli) cells and used directly after lyophilization. Several organic solvents and buffer amounts were screened for the reduction of the two substrates β‐carboline harmane and 1‐methyl‐3,4‐dihydroisoquinoline to the corresponding amines. Cyclopentyl methyl ether (CPME) proved to be the best solvent choice for the envisaged reduction. In addition, CPME is currently referred to as an environmentally benign solvent. Optimized reaction conditions were applied to 20 mM of the hardly water soluble substrates, leading to good conversions (up to 96%) and excellent enantiomeric excesses (>99%) in the best cases. The use of micro‐aqueous reaction systems opens the way to further applications of IREDs with hardly water soluble substrates.
Product selectivity and yield in chemical reactions can be limited by side product formation or low conversions due to equilibrium limitations. Extractive reaction systems (ERS) employ an in situ liquid‐liquid extraction that separates the product from the reaction phase to overcome these difficulties. The design of ERS requires a broad knowledge of the discipline of process intensification and extraction and reaction engineering. Furthermore, specific knowledge about the interplay of reaction and extraction phenomena is unique to ERS. This review gives an overview of the design of ERS and enables their application to any suitable reaction. 相似文献