A novel β-glucosidase was purified from pumpkin (Cucurbita moschata) seed by anion exchange chromatography and gel permeation chromatography, and its molecular mass was determined to be 42.8 kDa by gel permeation chromatography. The heterodimeric structure consisting of two subunits, free from disulfide bonds, was determined by native-PAGE analysis followed by zymography. The enzyme was maximally active at pH 4.0 and 70°C, and V_max, K_m, and k_cat values were 0.078 units mg⁻¹ protein, 2.22 mM, and 13.29 min⁻¹, respectively, employing p-nitrophenyl-β-d-glucopyranoside as the substrate. The high content of glycine determined by amino acid analysis implies that the enzyme possesses flexible conformations and interacts with cell membranes and walls in nature. Circular dichroism studies revealed that the high stability of the enzyme within the pH range of 2.0–10.0 is due to its reversible pH-responsive characteristics for α-helix–antiparallel β-sheet interconversion.

用阴离子交换色谱法和凝胶渗透色谱法从南瓜（Cucurbita moschata）种子中纯化出一种新型β-葡萄糖苷酶，凝胶渗透色谱法测定其分子量为42.8 kDa。由两个亚基组成的异二聚体结构，不含二硫键，通过天然 PAGE 分析和酶谱确定。该酶在 pH 4.0 和 70°C 时具有最大活性，V _max、K _m和k _cat值分别为 0.078 单位 mg ^-1蛋白质、2.22 mM 和 13.29 min ^-1，使用对硝基苯基-β- d-吡喃葡萄糖苷作为底物。通过氨基酸分析确定的高含量甘氨酸意味着该酶具有灵活的构象，并在自然界中与细胞膜和细胞壁相互作用。圆二色性研究表明，该酶在 2.0-10.0 的 pH 范围内的高稳定性是由于其对 α-螺旋-反平行 β-折叠相互转换具有可逆的 pH 响应特性。