Design of fusion enzymes for biocatalytic applications in aqueous and non-aqueous media

Biocatalytic cascades play a fundamental role in sustainable chemical synthesis. Fusion enzymes are one of the powerful toolboxes to enable the tailored combination of multiple enzymes for efficient cooperative cascades. Especially, this approach offers a substantial potential for the practical application of cofactor-dependent oxidoreductases by forming cofactor self-sufficient cascades. Adequate cofactor recycling while keeping the oxidized/reduced cofactor in a confined microenvironment benefits from the fusion fashion and makes the use of oxidoreductases in harsh non-aqueous media practical. In this mini-review, we have summarized the application of various fusion enzymes in aqueous and non-aqueous media with a focus on the discussion of linker design within oxidoreductases. The design and properties of the reported linkers have been reviewed in detail. Besides, the substrate loadings in these studies have been listed to showcase one of the key limitations (low solubility of hydrophobic substrates) of aqueous biocatalysis when it comes to efficiency and economic feasibility. Therefore, a straightforward strategy of applying non-aqueous media has been briefly discussed while the potential of using the fusion oxidoreductase of interest in organic media was highlighted. Copyright © 2022 Ma, Zhang, Vernet and Kara

Crosslinked Aggregates of Fusion Enzymes in Microaqueous Organic Media

Baeyer-Villiger monooxygenases (BVMOs) are attractive for selectively oxidizing various ketones using oxygen into valuable esters and lactones. However, the application of BVMOs is restrained by cofactor dependency and enzyme instability combined with water-related downsides such as low substrate loading, low oxygen capacity, and water-induced side reactions. Herein, we described a redox-neutral linear cascade with in-situ cofactor regeneration catalyzed by fused alcohol dehydrogenase and cyclohexanone monooxygenase in aqueous and microaqueous organic media. The cascade conditions have been optimized regarding substrate concentrations as well as the amounts of enzymes and cofactors with the Design of Experiments (DoE). The carrier-free immobilization technique, crosslinked enzyme aggregates (CLEAs), was applied to fusion enzymes. The resultant fusion CLEAs were proven to function in microaqueous organic systems, in which the enzyme ratios, water contents (0.5–5 vol. %), and stability have been systematically studied. The fusion CLEAs showed promising operational (up to 5 cycles) and storage stability

Growth differentiation factor 15 in patients with congenital dyserythropoietic anaemia (CDA) type II

Contains fulltext : 95939.pdf (publisher's version ) (Closed access)Congenital dyserythropoietic anaemias (CDAs) are heterogeneous, hereditary disorders hallmarked by ineffective erythropoiesis and tissue iron overload. Growth differentiation factor 15 (GDF15) was suggested to mediate iron overload in iron-loading anaemias, such as the thalassaemias and CDAI by suppressing hepcidin, the key regulator of iron absorption. Here, we show that serum GDF15 concentrations are elevated in subjects with CDAI and CDAII. Despite similar disease characteristics, CDAI patients present with significantly higher GDF15 concentrations compared to CDAII patients. Hepcidin concentrations are inappropriately low in CDAII patients considering the severe hepatic iron overload associated with this disorder. GDF15 significantly correlates with the degree of anaemia (Hb), the response of erythropoiesis (reticulocyte index) as well as with iron availability in the serum (transferrin saturation). The observation that GDF15 is elevated in CDAII patients is consistent with the proposal that GDF15 is among the erythroid factors down-regulating hepcidin and contributing to iron overload in conditions of dyserythropoiesis