Enhanced stability of a rumen-derived xylanase using SpyTag/SpyCatcher cyclization
| Autor: | Huan Wang, Jia-Wen Cao, Xiao-Bao Sun, Ye-Bo Zhou, Qian Wang, De-Ying Gao, Guo-Ying Qian, Jiakun Wang, Kai-Yue Wu, Yang-Nan Li |
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| Rok vydání: | 2019 |
| Předmět: |
0106 biological sciences
Circular dichroism Rumen Physiology Stereochemistry Lignocellulosic biomass Protein Engineering 01 natural sciences Applied Microbiology and Biotechnology Catalysis 03 medical and health sciences 010608 biotechnology Enzyme Stability Animals Glycoside hydrolase Thermostability chemistry.chemical_classification 0303 health sciences Endo-1 4-beta Xylanases Sheep 030306 microbiology Circular Dichroism General Medicine Hydrogen-Ion Concentration Fosmid Enzyme chemistry Cyclization Xylanase Thermodynamics Biotechnology |
| Zdroj: | World journal of microbiologybiotechnology. 36(2) |
| ISSN: | 1573-0972 |
| Popis: | Microbiota from herbivore rumen is of great interest for mining glycoside hydrolases for lignocellulosic biomass biorefinement. We previously isolated a highly active but poorly thermostable xylanase (LXY) from a rumen fluid fosmid library of Hu sheep, a local high-reproductive species in China. In this study, we used a universal enzyme-engineering strategy called SpyTag/SpyCatcher molecular cyclization to improve LXY stability via isopeptide-bond-mediated ligation. Both linear and cyclized LXY (L- and C-LXY, respectively) shared similar patterns of optimal pH and temperature, pH stability, and kinetic constants (km and Vmax). However, the C-LXY showed enhanced thermostability, ion stability, and resilience to aggregation and freeze-thaw treatment than L-LXY, without compromise of its catalytic efficiency. Circular dichroism and intrinsic and 8-anilino-1-naphthalenesulfonic acid-binding fluorescence analysis indicated that the cyclized enzyme was more capable of maintaining its secondary and tertiary structures than the linear enzyme. Taken together, these results promote the cyclized enzyme for potential applications in the feed, food, paper pulp, and bioenergy industries. |
| Databáze: | OpenAIRE |
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