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Stability Enhancement of Target Enzymes via Tyrosinase-Mediated Site-Specific Polysaccharide Coating

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Abstract

Enzymes are widely used in industrial and pharmaceutical applications, but their activities often decrease rapidly under harsh environmental conditions such as heat, organic solvents, and dehydration. In this study, a new method for enzyme coating with polysaccharide using a rapid tyrosinase-mediated crosslinking reaction was developed. When tyrosinase reacts with a monophenol-containing biopolymer such as polysaccharide, it forms a covalent crosslink between the biopolymer and the enzyme. This crosslinking reaction create a rigid polysaccharide-coated enzyme (PCE) that protects the enzyme from harsh environmental conditions, that leads to improve the enzyme stability. To demonstrate the concept, trypsin (TR), a model enzyme with a positively charged surface, was used. Tyramine conjugated alginate polymer (AlgT), a negatively charged biocompatible polysaccharide, was used to coat TR. The AlgT was subsequently used to coat TR, forming an AlgT-TR complex. We characterized the PCE using particle size, surface charge (zeta potential), optimal pH shift, etc. Afterwards, we compared the enzyme kinetics of AlgT-TR and uncoated TR (free-TR). The AlgT-TR showed a higher activity and higher heat, storage, and water-miscible organic solvent stabilities than the free-TR. The AlgT coating method was efficient and effective to increase the thermal stability of not only TR, but also hydrolases with neutral to negative surface charges, such as elastase, subtilisin, and chymotrypsin. These results suggest that the tyrosinase-mediated crosslinking reaction is a very promising and general coating method for improving the stability of enzymes with positive surface charge, but the opposite case would be also possible.

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Acknowledgements

This research was supported by Korea Initiative for fostering University of Research and Innovation Program of the National Research Foundation (NRF) funded by the Korean government (MSIT) (No. NRF-2020M3H1A1073304).

This work was also supported by the Korea Planning & Evaluation of Industrial Technology (20024336) funded by the Ministry of Trade, Industry & Energy (MOTTE, Republic of Korea).

This paper is based on the doctoral dissertation [37] submitted by Dr. Hyun Kim to Seoul National University in August 2022.

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Authors and Affiliations

  1. School of Chemical and Biological Engineering, Seoul National University, Seoul, 08826, Korea

    Hyun Kim, Uk-Jae Lee, Gyu-Min Lim, Jeongchan Lee, Hanbit Song, Nathaniel S. Hwang & Byung-Gee Kim

  2. Bio-MAX/N-Bio, Seoul National University, Seoul, 08826, Korea

    Hyun Kim, Uk-Jae Lee, Jeongchan Lee, Nathaniel S. Hwang & Byung-Gee Kim

  3. Interdisciplinary Program for Biochemical Engineering and Biotechnology, Seoul National University, Seoul, 08826, Korea

    Jin-Young Kim, Nathaniel S. Hwang & Byung-Gee Kim

  4. Institute for Sustainable Development (ISD), Seoul National University, Seoul, 08826, Korea

    Byung-Gee Kim

  5. R&D Center, EGC Therapeutics, Inc., Seoul, 08826, Korea

    Eun-jung Kim & Byung-Gee Kim

  6. Department of Chemical and Biological Engineering, Korea University, Seoul, 02841, Korea

    Jungbae Kim

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  1. Hyun Kim
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Kim, H., Lee, UJ., Lim, GM. et al. Stability Enhancement of Target Enzymes via Tyrosinase-Mediated Site-Specific Polysaccharide Coating. Biotechnol Bioproc E 28, 862–873 (2023). https://doi.org/10.1007/s12257-023-0190-5

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