【摘要】 果胶酸裂解酶（pectate lyases, Pel）在果胶降解中发挥重要作用，广泛应用于食品工业、纺织品加工等领域。本研究克隆并异源表达了1个来源于湖羊（Ovis aries）瘤胃微生物的果胶酸裂解酶基因IDSPL1-20 （GenBank No. PP975428），并对重组蛋白r IDSPL1-20的活性和酶学性质进行研究。结果表明，rIDSPL1-20的最适温度、pH分别为50℃、pH 10.0。该酶的热稳定性较差，但具有较强的耐碱能力，在pH 9.0～11.0下处理1 h残余活性仍大于80%。反应体系中添加0.25～2 mmol/L Ca²⁺能显著提升rIDSPL1-20的催化活性（P<0.05）。多底物分析显示，rIDSPL1-20降解聚半乳糖醛酸（polygalacturonic acid, PGA）和60%酯化果胶（60%DE pectin）的最大反应速度V_max分别为（316.25±34.09）和（105.55±3.72）μmol/（min·mg），降解产物主要为不饱和三聚半乳糖醛酸（unsaturated trigalacturonate, uG3）与不饱和二聚半乳糖醛酸（unsaturated digalacturonate, uG2），属于内切果胶酸裂解酶（endo-pectate lyase, EC 4.2.2.2）。此外，rIDSPL1-20能有效提升Cellic^?CTec3 HS对于花生（Arachis hypogaea）秸秆的降解效率，协同催化2和4 h后，总还原糖产量分别为（0.50±0.07）和（0.57±0.14） mg/mL。本研究为研发新型生物质降解酶建立基础。更多还原

【Abstract】 Pectate lyases（Pel） play critical roles in pectin degradation and hold significant biotechnological promise across diverse application in food and textile industries. In this study, a novel pectate lyase gene derived from Hu sheep（Ovis aries） rumen microbiota, IDSPL1-20（GenBank No. PP975428）, was cloned and heterologously expressed. The enzyme activity and biochemical characterizations of recombinant rIDSPL1-20were determined. The results showed that the optimum temperature and pH of rIDSPL1-20 were 50 ℃ and pH 10.0, respectively. The rIDSPL1-20 exhibited poor thermal stability but excellent alkaline-tolerance. The residual activity maintained over 80% after treatment at pH 9.0～11.0 for 1 h. The rIDSPL1-20 was activated by the presence of 0.25～2 mmol/L Ca²⁺（P<0.05）. Multi-substrate specificities determination indicated that the rIDSPL1-20 showed V_max values of（316.25±34.09） and（105.55±3.72） μmol/（min·mg） against polygalacturonic acid（PGA） and 60% esterified pectin（60% DE pectin）. The rIDSPL1-20 mainly released unsaturated trigalacturonate（uG3） and unsaturated digalacturonate（uG2） from PGA and 60% DE pectin as the predominant products, indicating that the enzyme probably functions as a typical endo-pectate lyase（EC4.2.2.2）. In addition, the degradation efficiency of Cellic^? CTec3 HS for peanut（Arachis hypogaea）straw was dramatically boosted by r IDSPL1-20（P<0.05）. After synergistic reaction for 2 and 4 h, the total reducing sugars yielded（0.50±0.07） and（0.57±0.14） mg/mL. This study laid the groundwork for the development of novel biomass-degrading enzymes.更多还原