To develop new cellulases for efficient utilization of the lignocellulose, an endoglucanase (CoCel5A) gene from Colletotrichum orchidophilum was synthesized and a recombinant Pichia pastoris GS115/pPIC9K/cocel5A was constructed for secretory expression of CoCel5A. After purification, the protein CoCel5A was biochemically characterized. The endoglucanase CoCel5A exhibited the optimal activity at 55–75 °C and high thermostability (about 85% residual activity) at the temperature of 55 °C after incubation for 3 h. The highest activity of CoCel5A was detected when 100 mM citric acid buffer (pH 4.0–5.0) was used and excellent pH stability (up to 95% residual activity) was observed after incubation in 100 mM citric acid buffer (pH 3.0–6.0) at 4 °C for 24 h. Carboxymethyl cellulose sodium salt (n = approx. 500) (CMC) and β-d-glucan were the best substrates for CoCel5A among the tested substrates. The kinetic parameters V_max, K_m, and K_cat/K_m values against CMC were 290.70 U/mg, 2.65 mg/mL, and 75.67 mL/mg/s, respectively; and 228.31 U/mg, 2.06 mg/mL, and 76.45 mL/mg/s against β-d-glucan, respectively, suggesting that CoCel5A has high affinity and catalytic efficiency. These properties supported the potential application of CoCel5A in biotechnological and environmental fields.

为了开发新的纤维素酶以有效利用木质纤维素，合成了来自Colletotrichum orchidophilum 的内切葡聚糖酶 (CoCel5A) 基因和重组毕赤酵母构建 GS115/pPIC9K/cocel5A 用于 CoCel5A 的分泌表达。纯化后，蛋白质 CoCel5A 被生化表征。内切葡聚糖酶 CoCel5A 在 55-75°C 下表现出最佳活性，在 55°C 温育 3 小时后表现出较高的热稳定性（约 85% 的残留活性）。当使用 100 mM 柠檬酸缓冲液（pH 4.0-5.0）时检测到 CoCel5A 的最高活性，并且在 100 mM 柠檬酸缓冲液（pH 3.0-6.0）中孵育后观察到优异的 pH 稳定性（高达 95% 残留活性） 4°C 24 小时。羧甲基纤维素钠盐 ( n  = 约 500) (CMC) 和β - d-葡聚糖是测试底物中 CoCel5A 的最佳底物。的动力学参数V_最大、K _m和K _cat / K _m值对 CMC 分别为 290.70 U/mg、2.65 mg/mL 和 75.67 mL/mg/s；和 228.31 U/mg、2.06 mg/mL 和 76.45 mL/mg/s 对β - d -葡聚糖，表明 CoCel5A 具有高亲和力和催化效率。这些特性支持 CoCel5A 在生物技术和环境领域的潜在应用。