Institute of Agricultural Products Processing and Nuclear Agriculture Technology Research, Hubei Academy of Agricultural Sciences, Wuhan 430064, P.R. China.
College of Biology and Pharmacy, Three Gorges University, Yichang 443002, P.R. China.
J Microbiol Biotechnol. 2022 Jun 28;32(6):749-760. doi: 10.4014/jmb.2201.01022. Epub 2022 May 16.
α-Galactosidase is a debranching enzyme widely used in the food, feed, paper, and pharmaceuticals industries and plays an important role in hemicellulose degradation. Here, T26, an aerobic bacterial strain with thermostable α-galactosidase activity, was isolated from laboratory-preserved lignocellulolytic microbial consortium TMC7, and identified as . The α-galactosidase, called T26GAL and derived from the T26 culture supernatant, exhibited a maximum enzyme activity of 0.4976 IU/ml when cultured at 60°C and 180 rpm for 2 days. Bioinformatics analysis revealed that the α-galactosidase T26GAL belongs to the GH36 family. Subsequently, the pET-26 vector was used for the heterologous expression of the T26 α-galactosidase gene in BL21 (DE3). The optimum pH for α-galactosidase T26GAL was determined to be 8.0, while the optimum temperature was 60°C. In addition, T26GAL demonstrated a remarkable thermostability with more than 93% enzyme activity, even at a high temperature of 90°C. Furthermore, Ca and Mg promoted the activity of T26GAL while Zn and Cu inhibited it. The substrate specificity studies revealed that T26GAL efficiently degraded raffinose, stachyose, and guar gum, but not locust bean gum. This study thus facilitated the discovery of an effective heat-resistant α-galactosidase with potent industrial application. Meanwhile, as part of our research on lignocellulose degradation by a microbial consortium, the present work provides an important basis for encouraging further investigation into this enzyme complex.
α-半乳糖苷酶是一种广泛应用于食品、饲料、造纸和制药行业的支链酶，在半纤维素降解中发挥着重要作用。本研究从实验室保存的木质纤维素降解微生物联合体 TMC7 中分离出一株具有耐热 α-半乳糖苷酶活性的需氧细菌 T26，并鉴定为. 从 T26 培养上清液中提取的 α-半乳糖苷酶 T26GAL 在 60°C、180rpm 培养 2 天时表现出最大酶活 0.4976IU/ml。生物信息学分析表明，T26GAL 属于 GH36 家族。随后，使用 pET-26 载体在 BL21（DE3）中异源表达 T26 α-半乳糖苷酶基因。α-半乳糖苷酶 T26GAL 的最适 pH 为 8.0，最适温度为 60°C。此外，T26GAL 具有显著的热稳定性，在 90°C 的高温下仍保持超过 93%的酶活。此外，Ca 和 Mg 促进了 T26GAL 的活性，而 Zn 和 Cu 则抑制了它。底物特异性研究表明，T26GAL 能有效降解棉子糖、水苏糖和瓜尔胶，但不能降解槐豆胶。因此，本研究发现了一种有效的耐热 α-半乳糖苷酶，具有潜在的工业应用价值。同时，作为我们对微生物联合体降解木质纤维素研究的一部分，本工作为进一步研究该酶复合物提供了重要依据。
