热稳定性
合理设计
化学
生物催化
突变体
生物化学
食品科学
酶
生物
催化作用
遗传学
基因
离子液体
作者
Wenli Zhang,Hu Ren,Xinxiu Wang,Quanyu Dai,Xiaoyong Liu,Dawei Ni,Yingying Zhu,Wei Xu,Wanmeng Mu
标识
DOI:10.1016/j.ijbiomac.2023.126919
摘要
Currently, alginate oligosaccharides (AOS) become attractive due to their excellent physiological effects. AOS has been widely used in food, pharmaceutical, and cosmetic industries. Generally, AOS can be produced from alginate using alginate lyase (ALyase) as the biocatalyst. However, most ALyase display poor thermostability. In this study, a thermostable ALyase from Paenibacillus sp. YN15 (Payn ALyase) was characterized. It belonged to the polysaccharide lyase (PL) 31 family and displayed poly β-D-mannuronate (Poly M) preference. Under the optimum condition (pH 8.0, 55 °C, 50 mM NaCl), it exhibited maximum activity of 90.3 U/mg and efficiently degraded alginate into monosaccharides and AOS with polymerization (DP) of 2-4. Payn ALyase was relatively stable at 55 °C, but the thermostability dropped rapidly at higher temperatures. To further improve its thermostability, rational design mutagenesis was carried out based on a combination of FireProt, Consensus Finder, and PROSS analysis. Finally, a triple-point mutant K71P/Y129G/S213G was constructed. The optimum temperature was increased from 55 to 70 °C, and the Tm was increased from 62.7 to 64.1 °C. The residual activity after 30 min incubation at 65 °C was enhanced from 36.0 % to 83.3 %. This study provided a promising ALyase mutant for AOS industrial production.
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