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- Title
定向引入 N‐糖基化位点促进芳基醇氧化酶 热稳定性及底物亲和力.
- Authors
曹查; 朱作华; 龚文兵; 周映君; 谢纯良; 彭源德
- Abstract
Aryl‐alcohol oxidase plays a crucial role in lignin degradation,and its enzymatic properties are influenced by N‐glycosylation modification. This study aimed to investigate the N‐glycosylation of Pleurotus eryngii aryl‐alcohol oxidase in order to enhance its thermal stability and substrate affinity. In this study,the expression system of Pichia GS115 and site‐directed mutagenesis were used to construct the expressions of six aryl‐alcohol oxidase mutant proteins. The purified wild‐type and mutant enzymes were also analyzed for enzymatic properties and stability. The results indicated that the N‐glycosylation mutations at sites N89 and N249 of aryl-alcohol oxidase caused reduced optimum temperature and enzymatic thermal stability at 70 ℃. However,introducing a new glycosylation site through mutagenesis did not affect the optimum pH but significantly improved both the optimum temperature and thermal stability at 70 ℃ compared to the wild‐type enzyme. When using p‐coumaric alcohol as a substrate,the mutant [AAO(F‐X‐N‐X‐T)] exhibited the highest substrate affinity. N‐gly‐cosylation primarily affects the thermal stability of aryl-alcohol oxidase,with glycosylation at sites N89 and N249 playing a critical role in enzyme stability. The introduction of N‐glycosylation site [AAO(F‐X‐N‐X‐T)] results in aryl‐alcohol oxidase with enhanced activity and stability.
- Publication
Food Research & Development, 2024, Vol 45, Issue 7, p165
- ISSN
1005-6521
- Publication type
Article
- DOI
10.12161/j.issn.1005‐6521.2024.07.023