Valency engineering of monomeric enzymes for self-assembling biocatalytic hydrogels

Bitterwolf, Patrick; Gallus, Sabrina; Peschke, Theo; Mittmann, Esther; Oelschlaeger, Claude; Willenbacher, Norbert; Rabe, Kersten S.; Niemeyer, Christof M.

doi:10.1039/c9sc04074a

Cited by 37 publications

(45 citation statements)

References 29 publications

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“…Microrheological experiments were carried out to confirm the polymeric nature of the hydrogels and to quantitatively determine the degree of crosslinking in the various H1-H4 materials in order to identify the most stable gel for the subsequent flow experiments (Figure 4). All hydrogels were obtained as monolithic, elastic materials with rheological properties comparable to hydrogels prepared from different enzymes, as reported previously by our group [44,46]. Figure 4a shows the variation of mean square-displacements (MSDs) as a function of lag time for polystyrene particles with a diameter of 200 nm dispersed in the hydrogel H1.…”

Section: Resultsmentioning

confidence: 56%

“…Therefore, our study does not only expand the scope of all-enzyme hydrogels by adding a new enzyme with high application relevance to the toolbox. The present work also suggests that our approach holds great potential for the establishment of advanced microfluidic manufacturing processes that include numbering up of chips to improve productivity [46] or the implementation of biocatalytic and chemoenzymatic cascades in machine-assisted compartmentalized production processes [32].…”

Section: Discussionmentioning

confidence: 79%

“…We have recently demonstrated the construction of self-assembling all-enzyme hydrogels constructed from ketoreductases and glucose dehydrogenases, which were genetically fused with either the SpyTag (ST) peptide or the SpyCatcher (SC) protein, respectively [44][45][46]. This highly efficient autocatalytic bioconjugation system is based on the rapid formation of a covalent isopeptide bond through the SpyTag-SpyCatcher complex that occurs under physiological conditions [47][48][49][50].…”

Section: Introductionmentioning

confidence: 99%

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A Phenolic Acid Decarboxylase-Based All-Enzyme Hydrogel for Flow Reactor Technology

et al. 2019

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Carrier-free enzyme immobilization techniques are an important development in the field of efficient and streamlined continuous synthetic processes using microreactors. Here, the use of monolithic, self-assembling all-enzyme hydrogels is expanded to phenolic acid decarboxylases. This provides access to the continuous flow production of p-hydroxystyrene from p-coumaric acid for more than 10 h with conversions ≥98% and space time yields of 57.7 g·(d·L)−1. Furthermore, modulation of the degree of crosslinking in the hydrogels resulted in a defined variation of the rheological behavior in terms of elasticity and mesh size of the corresponding materials. This work is addressing the demand of sustainable strategies for defunctionalization of renewable feedstocks.

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Section: Resultsmentioning

confidence: 56%

Section: Discussionmentioning

confidence: 79%

Section: Introductionmentioning

confidence: 99%

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A Phenolic Acid Decarboxylase-Based All-Enzyme Hydrogel for Flow Reactor Technology

et al. 2019

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“…147 First results showed high conversion rates and stereoselectivities as well as high space-time yields. 121,147 In addition, the in situ generation of pure enzyme hydrogels easily enables an increase in protein loading for microfluidic bioreactors. It is also Fig.…”

Section: Carrier-free Methodsmentioning

confidence: 99%

“…Two major challenges need to be overcome for a possible industrial application of all-enzyme hydrogels: firstly, this approach is limited to multimeric enzymes (as a recent survey of the PDB database pointed out that only ∼10% of the enzymes are homotetramers), 121 and secondly, a slight decrease in the enzyme activity is observed (probably due to mass transport limitations). 120 Fortunately, this challenge could be solved by enzyme engineering.…”

Section: Carrier-free Methodsmentioning

confidence: 99%

The rise of continuous flow biocatalysis – fundamentals, very recent developments and future perspectives

2020

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Flow‐Induced Microfluidic Assembly for Advanced Biocatalysis Materials

Lemke,

Schneider,

Kunz

et al. 2024

Adv Funct Materials

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Exploring the potential of microfluidic systems, this study presents a groundbreaking approach harnessing energy in microfluidic flows within a purpose‐built microreactor, enabling precise deposition of functional biomaterials. Upon optimizing reactor dimensions and integrating it into a microfluidic system, sequentially flow‐induced deposition of DNA hydrogels and transformation into DNA‐protein hybrid materials with SpyTag/SpyCatcher technology is investigated. However, limited functionalization rates restrict its viability for targeted biocatalytic processes. Therefore, the direct deposition of a phenolic acid decarboxylase is investigated, which is efficiently deposited but shows limited biocatalytic performance due to shear‐induced denaturation. This challenge is overcome by a two‐step immobilization process, resulting in microfluidic bioreactors demonstrating initial high space‐time yields of up to 7000 g L−1 d−1, but whose process stability proves unsatisfactory. However, by exploiting the principle of flow‐induced deposition to immobilize recombinant E. coli cells as functional living materials overexpressing biocatalytically relevant enzymes, bioreactors are produced that show equally high space‐time yields in continuous whole‐cell catalysis which remain constant over periods of up to 10 days. The insights gained offer optimization strategies for advanced functional materials and innovative reactor systems holding promise for applications in fundamental materials science, biosensing, and scalable production of microreactors for biocatalysis and bioremediation.

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Valency engineering of monomeric enzymes for self-assembling biocatalytic hydrogels

Abstract: All-enzyme hydrogels are efficient reagents for continuous flow biocatalysis.

Cited by 37 publications

References 29 publications

A Phenolic Acid Decarboxylase-Based All-Enzyme Hydrogel for Flow Reactor Technology

A Phenolic Acid Decarboxylase-Based All-Enzyme Hydrogel for Flow Reactor Technology

The rise of continuous flow biocatalysis – fundamentals, very recent developments and future perspectives

Flow‐Induced Microfluidic Assembly for Advanced Biocatalysis Materials

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