Enzyme cofactors play a major role in biocatalysis, as many enzymes require them to catalyze highly valuable reactions organic synthesis. However, the cofactor recycling is often hurdle implement at industrial level. The fabrication of heterogeneous biocatalysts co-immobilizing phosphorylated (PLP, FAD+ , and NAD+ ) onto same solid material reported perform chemical without exogeneous addition aqueous media. In these self-sufficient biocatalysts, immobilized are catalytically active...

Abstract The mechanisms that underlie the regulation of enzymatic reactions by biomolecular condensates and how they scale with compartment size remain poorly understood. Here we use intrinsically disordered domains as building blocks to generate programmable NADH-oxidase (NOX) different sizes spanning from nanometers microns. These domains, derived three distinct RNA-binding proteins, each possessing net charge, result in formation characterized a comparable high local concentration enzyme...

We expanded the application of self-sufficient heterogeneous biocatalysts containing coimmobilized ω-transaminases and pyridoxal 5′-phosphate (PLP) to efficiently operate packed-bed reactors in continuous flow. Using a ω-transaminase from Halomonas elongata with PLP onto porous methacrylate-based carriers coated polyethylenimine, we operated reactor continuously for up 50 column volumes at 1.45 mL × min–1 enantioselective deamination model amines (α-methylbenzyl amine), yielding >90%...

<bold>Flow biocatalysis: where to start?</bold> This tutorial review aims guide and inspire new-comers the field boost potential of flow biocatalysis.

The development of cell-free and self-sufficient biocatalytic systems represents an emerging approach to address more complex synthetic schemes under nonphysiological conditions. Herein, we report the a heterogeneous biocatalyst for synthesis chiral alcohols without need add exogenous cofactor. In this work, NADPH-dependent ketoreductase was primarily stabilized further co-immobilized with NADPH catalyze asymmetric reductions addition As result, immobilized cofactor is accessible, thus, it...

As an alternative to the traditional chemical synthesis or <italic>in vivo</italic> production of <sc>l</sc>-pipecolic acid, we have developed two <italic>ex strategies using purified and immobilised enzymes for this key building block.

The continuous synthesis of valuable nucleoside drugs was achieved in up to 99 % conversion by using a novel halotolerant purine phosphorylase from Halomonas elongata (HePNP). HePNP showed an unprecedented tolerance DMSO, usually required for substrate solubility, and could be immobilized on agarose microbeads through disulfide bonds, via genetically fused Cystag. This covalent yet reversible binding chemistry showcased the reusability second round enzyme immobilization with high...

Abstract Flow biocatalysis has emerged as an empowering tool to boost the potential of enzymatic reactions towards more automatized, sustainable, and generally efficient synthetic processes. In last fifteen years, increasing number biocatalytic transformations carried out in continuous flow exemplified benefits that this technology can bring incorporate into industrial operations. This perspective aims capture a nutshell available methodologies for well discuss current limitations future...

Cell-free biocatalysis is gaining momentum in producing value-added chemicals, particularly stepwise reaction cascades. However, the stability of enzyme cascades industrial settings often compromised when free enzymes are involved. In this study, we have developed a stable multifunctional heterogeneous biocatalyst coimmobilizing five on microparticles to transform 1,ω-diols into 1,ω-hydroxy acids. We improved operational efficiency and by fine-tuning loading spatial organization. Stability...

Understanding how the immobilization of enzymes on solid carriers affects their performance is paramount for design highly efficient heterogeneous biocatalysts. An supply substrates onto phase one main challenges to maximize activity immobilized enzymes. Herein, we apply advanced single-particle analysis decipher optimal an NADH oxidase (NOX) whose depends both O2 and concentrations. Carrier physicochemical properties its functionality along with enzyme distribution across carrier were...

Lignin has emerged as an attractive alternative in the search for more eco-friendly and less costly materials enzyme immobilization. In this work, terephthalic aldehyde-stabilization of lignin is carried out during its extraction to develop a series functionalized lignins with range reactive groups (epoxy, amine, aldehyde, metal chelates). This expands immobilization pool enzymes (carboxylase, dehydrogenase, transaminase) by different binding chemistries, affording yields 64-100 %. As proof...

Heterocyclic amines are a key structural motif for the synthesis of pharmaceuticals (e.g., antibiotics) as well pesticides and flavors. In this regard, imine reductases (IREDs) have recently emerged highly selective sustainable alternative asymmetric reductive amination reactions. Herein, we applied six IREDs, two which were newly identified, in reduction heterocyclic imines with either N, S, or O substitution at C-4. Since IREDs NADPH-dependent enzymes, commercially available, supported...

Abstract In this perspective article, we celebrate the accomplishments of female‐led research groups in biocatalysis. Through initiative, aim to showcase breadth and excellence women's increase their visibility within catalysis community. The authors wish emphasize that article represents only a small selection extraordinary women who have shaped field biocatalysis over time. Among them are scientists directly or significantly influenced inspired authors’ scientific journeys.

Fabrication of protein-based biomaterials is an arduous and time-consuming procedure with multiple steps. In this work, we describe a portable toolkit that integrates both cell-free protein synthesis (CFPS) immobilization in one pot just by mixing DNA, solid materials, CFPS system. We have constructed modular set plasmids fuse the N-terminus superfolded green fluorescent (sGFP) different peptide tags (poly(6X)Cys, poly(6X)His, poly(6X)Lys), which drive on tailored material (agarose beads...

Enzyme immobilization has become a key strategy to improve the stability and recycling of biocatalysts, resulting in greener more cost-efficient processes. The design immobilized catalysts is often focused only on strategy, binding chemistry between enzyme support, while less attention been paid physico-chemical properties material supports. Selecting best carrier for specific application may greatly influence performance biocatalytic reaction. Herein, we present comparative study two most...

Abstract l ‐Pipecolic Acid ( ‐PA) is a valuable building block for the synthesis of pharmaceuticals such as anesthetics and immunosuppressants. Thus, more efficient greener strategies are desired its production. Herein, we have applied previously engineered variant Lysine Cyclodeaminase from Streptomyces pristinaespiralis (e‐SpLCD) bioconversion ‐Lysine into ‐PA. The reaction can be performed by free e‐SpLCD reaching full conversion to 50 mM From biotechnological perspective, process...

Squalene-hopene cyclases are a highly valuable and attractive class of membrane-bound enzymes as sustainable biotechnological tools to produce aromas bioactive compounds at industrial scale. However, their application whole-cell biocatalysts suffer from the outer cell membrane acting diffusion barrier for hydrophobic substrate/product, while use purified leads dramatic loss stability. Here we present an unexplored strategy biocatalysis: squalene-hopene-cyclase spheroplasts. By removing...

Several chemoenzymatic routes have been explored for the preparation of cinacalcet, a calcimimetic agent. Transaminases (TAs) and ketoreductases (KREDs) turned out to be useful biocatalysts key optically active precursors. Thus, asymmetric amination 1-acetonaphthone yielded an enantiopure (R)-amine, which can alkylated in one step yield cinacalcet. Alternatively, bioreduction same ketone resulted (S)-alcohol, was easily converted into previous (R)-amine. In addition, reduction efficiently...

Abstract Fusion enzymes are attractive tools for facilitating the assembly of biocatalytic cascades chemical synthesis. This approach can offer great advantages cooperative redox that need constant supply a donor molecule. In this work, we have developed self‐sufficient bifunctional enzyme be coupled to transaminase‐catalyzed reactions efficient recycling amino (L‐alanine). By genetic fusion an alanine dehydrogenase (AlaDH) and formate (FDH), redox‐complementary system was applied recycle...

Abstract Immobilized enzymes have been widely exploited because they work as heterogeneous biocatalysts, allowing their recovery and reutilization easing the downstream processing once chemical reactions are completed. Unfortunately, we suffer a lack of analytical methods to characterize those biocatalysts at microscopic molecular levels with spatio‐temporal resolution, which limits design optimization. Single‐particle studies vital optimize performance immobilized in micro/nanoscopic...

We herein describe a bioinspired solid-phase assembly of multienzyme system scaffolded on an artificial cellulosome. An alcohol dehydrogenase and ω-transaminase were fused to cohesin dockerin domains drive their sequential ordered coimmobilization agarose porous microbeads. The resulting immobilized enzymatic cellulosome was characterized through quartz crystal microbalance with dissipation confocal laser scanning microscopy demonstrate that both enzymes interact each other physically...

Abstract Enzyme cofactors play a major role in biocatalysis, as many enzymes require them to catalyze highly valuable reactions organic synthesis. However, the cofactor recycling is often hurdle implement at industrial level. The fabrication of heterogeneous biocatalysts co‐immobilizing phosphorylated (PLP, FAD + , and NAD ) onto same solid material reported perform chemical without exogeneous addition aqueous media. In these self‐sufficient biocatalysts, immobilized are catalytically active...

A novel thymidine phosphorylase from H. elongata has been characterized, immobilized, and applied in a flow reactor. With this biocatalyst, four halogenated nucleoside analogues with anticancer antiviral properties were produced high yields.