Mahy, J.-P.

Biotechnol. Appl. Biochem. 2020, 67, 563-573, 10.1002/bab.1906

We report a new artificial hydrogenase made by covalent anchoring of the iron Knölker's complex to a xylanase S212C variant. This artificial metalloenzyme was found to be able to catalyze efficiently the transfer hydrogenation of the benchmark substrate trifluoroacetophenone by sodium formate in water, yielding the corresponding secondary alcohol as a racemic. The reaction proceeded more than threefold faster with the XlnS212CK biohybrid than with the Knölker's complex alone. In addition, efficient conversion of trifluoroacetophenone to its corresponding alcohol was reached within 60 H with XlnS212CK, whereas a ≈2.5-fold lower conversion was observed with Knölker's complex alone as catalyst. Moreover, the data were rationalized with a computational strategy suggesting the key factors of the selectivity. These results suggested that the Knölker's complex was most likely flexible and could experience free rotational reorientation within the active-site pocket of Xln A, allowing it to access the subsite pocket populated by trifluoroacetophenone.

Fasan, R.

Curr. Opin. Green Sustain. Chem. 2021, 31, 100494, 10.1016/j.cogsc.2021.100494

The direct functionalization of C–H bonds constitutes a powerful strategy to construct and diversify organic molecules. However, controlling the chemo- and site-selectivity of this transformation, particularly in complex molecular settings, represents a significant challenge. Metalloenzymes are ideal platforms for achieving catalyst-controlled selective C–H bond functionalization as their reactivities can be tuned by protein engineering and/or redesign of their cofactor environment. In this review, we highlight recent progress in the development of engineered and artificial metalloenzymes for C–H functionalization, with a focus on biocatalytic strategies for selective C–H oxyfunctionalization and halogenation as well as C–H amination and C–H carbene insertion via abiological nitrene and carbene transfer chemistries. Engineered heme and nonheme iron dependent enzymes have emerged as promising scaffolds for executing these transformations with high chemo-, regio-, and stereocontrol as well as tunable selectivity. These emerging systems and methodologies have expanded the toolbox of sustainable strategies for organic synthesis and created new opportunities for the generation of chiral building blocks, the late-stage C–H functionalization of complex molecules, and the total synthesis of natural products.

Yilmaz, F.

Process Biochem. 2018, 72, 71-78, 10.1016/j.procbio.2018.06.005

Carbonic anhydrase (carbonic dehydratase) (CA) is a metalloenzyme that contains zinc (Zn2+) ion in its active site. CA catalyzes the reversible conversion of carbon dioxide and water to bicarbonate and protons. Zn2+ ions, which are present in the active site of the enzyme, interact with the substrate molecules directly and cause catalytic effect. In this study, a nano-enzyme system was designed in aqueous solutions at room temperature and under nitrogen atmosphere to use the CA enzyme without any pre-treatment and deformation in its structure. The novel concept ANADOLUCA (AmiNo Acid (monomer) Decorated and Light Underpinning Conjugation Approach) was used for this process, nano CA enzyme of size 93 nm was synthesized. The activity of the synthesized nano CA was measured following the change in absorbance during the conversion of 4-nitrophenylacetate (NPA) to 4-nitrophenylate ion at 348 nm for a period of 10 min at 25 °C compared with free CA enzyme. Km and Vmax values for nano CA enzyme were found to be 0.442 mM and 1.6 × 10−3 mM min-1, respectively, whereas Km and Vmax values for free CA were found to be 0.471 mM and 1.5 × 10−3 mM min-1, respectively. In addition to these, the Zn2+ ion present in the active site of the nano CA enzyme was replaced by rodium metal. This nanorodium-substituted CA has been investigated as a new reductase enzyme for the stereoselective hydrogenation of olefins. Then, the Zn2+ ion in the active site of the nano CA enzyme was replaced with manganese metal to enhance the enzyme structure, thereby gaining characteristics of peroxidase. This newly synthesized nano manganese-substituted CA enzyme was investigated for its role as a peroxidase, which could be an alternative for hydrogen peroxidases.

Lin, Y.-W.

Biotechnol. Appl. Biochem. 2019, 10.1002/bab.1788

Environmental pollutants, such as industrial dyes and halophenols, are harmful to human health, which urgently demand degradation. Bioremediation has been shown to be a cost‐effective and ecofriendly approach. As reviewed herein, significant progress has been made in the last decade for biodegradation of both industrial dyes and halophenols, by engineering of native dye‐decolorizing peroxidases (DyPs) and dehaloperoxidases (DHPs), and by design of artificial heme enzymes in both native and de novo protein scaffolds. The catalytic efficiency of artificial DyPs and DHPs can be rationally designed comparable to or even beyond those of natural counterparts. The enzymes are on their way from laboratory to industry and will play more crucial roles in environmental protection toward a green future.

Berggren, G.

Curr. Opin. Green Sustain. Chem. 2021, 32, 100521, 10.1016/j.cogsc.2021.100521

Hydrogenases are gas processing redox enzymes central in hydrogen metabolism. The interdisciplinary nature of hydrogenase research is underscored by the development of “artificial maturation”, enabling the preparation of semi-synthetic hydrogenases through the incorporation of synthetic cofactors into a range of apo-hydrogenase hosts under in vitro and in vivo conditions. Herein, we discuss how the preparation of such semi-synthetic [FeFe]-hydrogenases has elucidated structural elements of the cofactor critical for catalysis and reactivity towards known inhibitors. It has also provided a convenient method for exploring the biodiversity of this enzyme family and thereby facilitated investigation of the role of the outer-coordination sphere in tuning the reactivity of the H-cluster. In parallel, hijacking the assembly line of the [FeFe]-hydrogenase through incorporation of synthetic precursors has provided detailed insight into the biosynthesis of the H-cluster. Moreover, it has allowed the preparation of Mn analogs of [Fe] hydrogenase.