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KMID : 1094720230280050862
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Biotechnology and Bioprocess Engineering
2023 Volume.28 No. 5 p.862 ~ p.873
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Stability Enhancement of Target Enzymes via Tyrosinase-Mediated Site-Specific Polysaccharide Coating
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Kim Hyun
Lee Uk-Jae
Lim Gyu-Min
Kim Jin-Young
Lee Jeong-Chan
Song Han-Bit
Kim Eun-Jung
Kim Jung-Bae
Hwang Nathaniel S.
Kim Byung-Gee
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Abstract
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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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KEYWORD
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enzyme stabilization, enzyme-polymer coating complex, tyrosinase, alginate, trypsin
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