An immobilized and highly stabilized self‐sufficient monooxygenase as biocatalyst for oxidative biotransformations

Abstract

BACKGROUND The requirement for expensive cofactors that must be efficiently recycled is one of the major factors hindering the wide implementation of industrial biocatalytic oxidation processes. In this research, a sustainable approach based on immobilized self‐sufficient Baeyer–Villiger monooxygenases is discussed. RESULTS A bifunctional biocatalyst composed of an NADPH‐dependent cyclohexanone monooxygenase (CHMO) fused to an NADP+‐accepting phosphite dehydrogenase (PTDH) catalyzes ϵ‐caprolactone synthesis from cyclohexanone, using phosphite as a cheap sacrificial substrate for cofactor regeneration. Several immobilized derivatives of the fused enzyme have been prepared with high immobilization yield (97%); the one obtained by affinity adsorption on Co‐IDA (Co: cobalt chelated; IDA: iminodiacetic acid) support has shown to be highly stable affording average yields of 80% after 18 reaction cycles. CONCLUSIONS The immobilized self‐sufficient monooxygenase has demonstrated to be able to perform Baeyer–Villiger oxidation with efficient cofactor recovery and biocatalyst recycling. The proposed biocatalytic process offers access to valuable molecules with high atom economy and limited waste generation. © 2017 Society of Chemical Industry

Ketoisophorone Synthesis with an Immobilized Alcohol Dehydrogenase

“…[44] In order to calculate the theoretical final specific activity of the immobilized derivate (U g À 1 support) the procedure explained below these lines was followed. For the M1, two pH values were tested: 8 and 9 (KPi, 1 M).…”

“…Due to the impossibility of measuring the actual suspension activity, most probably due to the presence of diffusional limitations, the final specific activity was calculated assuming that the retained activity (%) is maintained from the characterization study. [44] In order to calculate the theoretical final specific activity of the immobilized derivate (U g À 1 support) the procedure explained below these lines was followed. Attached units U g À 1 ð Þ ¼ Activity loaded U mL À 1 ð Þ À Activity supernatant U mL À 1 ð Þ Support loaded g ð Þ…”

Pepsin immobilization on biochar by adsorption and covalent binding, and its application for hydrolysis of bovine casein

“…Several methods have been used for enzyme immobilization, including adsorption and covalent bonding . The adsorption method is the simplest and most used method, and consists of the contact between the enzyme and the support, through weak bonds, such as Van der Waals forces, ionic interactions, physical adsorption or hydrophobic interactions between the enzyme constituents and the surface of the support material.…”

“…10 Several methods have been used for enzyme immobilization, including adsorption and covalent bonding. 11 The adsorption method is the simplest and most used method, and consists of the contact between the enzyme and the support, through weak bonds, such as Van der Waals forces, ionic interactions, physical adsorption or hydrophobic interactions between the enzyme constituents and the surface of the support material. Parameters such as pH, ionic strength and temperature must be controlled due to the types of forces involved in the process, because variations in these parameters may interfere with the adsorptive efficiency of the materials.…”

Efficient synthesis of the key chiral alcohol intermediate of Crizotinib using dual‐enzyme@CaHPO4 hybrid nanoflowers assembled by mimetic biomineralization

“…Compared with free enzyme, immobilized enzyme presents several advantages: first, immobilized enzyme can be easily fixed on the surface of carriers of various shapes such as granular, tubular, and membranous, which is easy to remove and facilitates continuous and repeated use. Moreover, improved tolerance to environment conditions can be observed after immobilization, resulting in enhanced stability and longer working cycle for immobilized enzymes.…”