Abstract

GH11 endo-xylanases, due to their inherent structural and biochemical properties, are the key to efficient bioconversion of lignocellulosic biomass into value-added products. A GH11 endo-xylanase (XynB) from Bacillus subtilis strain CAM 21 was cloned, over-expressed and purified (Mw∼24 kDa) using Ni-NTA affinity chromatography. XynB showed optimum activity at pH 7.0 and 50°C and was stable (>88%) in a broad range of pH (4–11). The apparent Km_, K_cat and K_cat/Km of XynB were 2.9 mg/ml, 1961.2/sec, and 675.62 ml/mg/sec, respectively using birchwood xylan as substrate. XynB was a classical endo-xylanase as it hydrolyzed birchwood xylan to xylo-oligosaccharides and not xylose. Kinetic stability of XynB at 45–53°C was between 43-182 min. Secondary structure analysis of XynB using far-UV CD spectroscopy revealed presence of 51.85% β strands and 2.64% α helix and was consistent with the homology modeling studies. XynB hydrolyzed the xylan extracted from agro-industrial wastes and fruit/vegetable peels by releasing up to 670 mg/g of reducing sugars. The xylan extracted from weeds (Ageratum conyzoides, Achyranthes aspera and Tridax procumbens) had characteristic signatures of hemicelluloses and after XynB hydrolysis showed cracks, peeling and release of up to 135.2 mg/g reducing sugars.

摘要

GH11 内切木聚糖酶由于其固有的结构和生化特性，是将木质纤维素生物质有效生物转化为增值产品的关键。来自枯草芽孢杆菌CAM 21 菌株的 GH11 内切木聚糖酶 (XynB)被克隆、过表达和纯化 (Mw~24 kDa)，使用 Ni-NTA 亲和层析。XynB 在 pH 7.0 和 50°C 下表现出最佳活性，并且在广泛的 pH (4-11) 范围内稳定 (>88%)。表观 Km _, K _cat和 K _catXynB 的 /Km 分别为 2.9 mg/ml、1961.2/sec 和 675.62 ml/mg/sec，使用桦木木聚糖作为底物。XynB 是一种经典的内切木聚糖酶，因为它将桦木木聚糖水解为低聚木糖而不是木糖。XynB 在 45-53°C 下的动力学稳定性在 43-182 分钟之间。使用远紫外 CD 光谱对 XynB 进行的二级结构分析显示存在 51.85% β 链和 2.64% α 螺旋，并且与同源建模研究一致。XynB 水解从农业工业废物和水果/蔬菜皮中提取的木聚糖，释放高达 670 mg/g 的还原糖。从杂草中提取的木聚糖（Ageratum conyzoides、Achyranthes aspera和Tridax procumbens) 具有半纤维素的特征特征，并且在 XynB 水解后显示裂纹、剥离和释放高达 135.2 mg/g 的还原糖。