Bypassing Biocatalytic Substrate Limitations in Oxidative Dearomatization Reactions by Transient Substrate Mimicking.

Enzymatic oxidative dearomatization is an efficient way to generate chiral molecules from simple arenes. One example is the flavin-dependent monooxygenase SorbC involved in sorbicillinoid biosynthesis. However, SorbC requires a long-chain keto substituent at its phenolic substrate, thus preventing its application beyond the synthesis of natural sorbicillinoids or close structural analogues. This work describes an approach to broaden the accessible product spectrum of SorbC by employing an ester functionality mimicking the natural substrate structure during enzymatic oxidation.

Hairy Particles with Immobilized Enzymes: Impact of Particle Topology on the Catalytic Activity.

Enzymes are described as ideal green biocatalysts because they are highly specific and selective. However, their practical application is hampered because of the low stability and missing reusability of free enzymes. One method to overcome these problems is the immobilization of enzymes onto carriers. Although numerous publications discuss different immobilization strategies, optimization of these carriers for the highest enzyme activity and loading capacity, enzyme selectivity, reusability, and reactor system configuration still remains a challenging task. In this contribution, we aim to address the role of the core-shell particle design with respect to their geometry as well as the polymer shell thickness on the immobilization of biomolecules. We discovered that spherical particles with a core diameter of 200 nm and intermediate shell thickness as well as platelet-like particles exhibited excellent results with a maximum immobilization yield of laccase from Trametes versicolor of up to 92% and an activity on the carrier material of 5.722 U/(g particle). Especially, the platelet-like particles offered a scalable and convenient alternative for the immobilization of laccase. Circular dichroism measurements proved that the secondary structure of the enzyme is not impaired by immobilization onto all kinds of carrier particles. Moreover, the immobilized laccase was successfully used for the decolorization of Cibacron blue P-3R in up to 18 cycles. Finally, particle separation was achieved via citrate-induced flocculation within 10 min. This detailed study contributes to the understanding of rational design of catalytically active hybrid materials and their effective performance at interfaces for applications in textile industry and environmental technologies.

Controlled and tunable design of polymer interface for immobilization of enzymes: does curvature matter?

Control and tuning of surface properties is indispensable for the programmed and rational design of materials. Particularly, polymeric brush-modified colloids can be used as carrier materials for enzyme immobilization. Although it is of prime importance to control the brush architecture, there is still a lack of systematic investigations concerning the impact of grafting density on the properties of the designed interface, as well as on the immobilization of biomolecules. In this work, we investigate the surface properties of polymer brushes with different grafting densities prepared using a "grafting from" approach on flat and on colloidal particle substrates by varying the density of initiator groups. In this way, we control and tune interfacial properties of the carrier material such as swelling, charge, adhesion as well as adsorption of laccase from Trametes versicolor on the grafted polyelectrolyte layer. We show that there is no direct transferability of the results received from planar to curved substrates regarding the swelling behavior in dependence on the grafting density. The maximum of swelling degree of PDMAEMA layers is achieved at 0.34 nm-2 and at 0.1 nm-2 grafting density for planar and curved particle substrates, respectively. The adhesion properties of the polymeric layer on both substrates are also strongly influenced by the grafting density, i.e. a decrease of the grafting density causes a transition from the adhesive to non-adhesive state. As proven by the cryo-TEM and AFM force distance measurements, an immobilization of laccase from Trametes versicolor causes a decrease of the polymer swelling and therefore leads to the changes in the surface morphology, charge and adhesion performance of final polymer-enzyme layer. Moreover, the higher effectiveness and activity of laccase were observed for the intermediate grafting densities which seem to be preferable over the maximum brush densities.

One-copper laccase-related enzyme from Marasmius sp.: purification, characterization and bleaching of textile dyes.

In the culture filtrate of a Marasmius sp. strain isolated in Indonesia during a screening for fungi with the ability to decolorize textile dyes, two laccase-related enzymes (laccase-related enzyme I and II) were detected. Laccase-related enzyme I was purified to homogeneity by ion exchange and hydrophobic interaction chromatography. The native enzyme was shown to have a molecular mass of 53 kDa, an N-terminal amino acid sequence characteristically seen in laccases and an isoelectric point of pH 3.8. The enzyme accepts typical laccase substrates including 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS), syringaldazine and guaiacol, but has no tyrosinase activity. The pH optimum is at pH 3.0 for ABTS and at 6.0 for syringaldazine and the enzyme is stable up to pH 10. The UV/vis spectrum of the laccase-related enzyme is non-typical for laccases and metal content analysis revealed that the enzyme contains only a single copper atom per enzyme molecule. This suggests that this enzyme could be related to the group of the so-called "white" laccases, however, no zinc or any other metal ion could be detected in this enzyme, suggesting that the enzyme is a unique laccase-related enzyme. Comparison of the bleaching activity of the whole fungus with that of the isolated laccase-related enzyme showed that this enzyme is the major bleaching enzyme produced by this Marasmius sp. strain and was able to bleach violet, red, orange and yellow dyes in addition to a number of blue dyes.