木聚糖酶Xyn11A与阿拉伯呋喃糖苷酶Abf62A协同水解麦秆木聚糖生物合成低聚木糖

doi:10.12211/2096-8280.2025-037

摘要/Abstract

摘要：

低聚木糖（XOSs）是一类由2-7个木糖分子通过β-1，4糖苷键连接而成的功能性寡糖，被公认为是具有益生元活性的可溶性膳食纤维。在温和条件下的酶催化从廉价生物质提取木聚糖制备高附加值低聚木糖引起了广泛关注。本研究从宇佐美曲霉中克隆了一种阿拉伯呋喃糖苷酶Abf62A基因，并实现其在毕赤酵母X33中的异源表达。利用木聚糖酶Xyn11A（200 U/g 底物）和阿拉伯呋喃糖苷酶Abf62A（20 U/g 底物）对高浓度麦秆木聚糖（100 g/L）进行协同酶水解，生成50.32 g/L低聚木糖，相较于木聚糖酶Xyn11A单酶水解（34.44 g/L 低聚木糖）展现了显著的协同效应。协同水解低聚木糖产物分析结果显示50.32 g/L低聚木糖中包含木二糖（31.71 g/L）、木三糖（15.92 g/L）、木四糖（1.65 g/L）和木五糖（1.04 g/L），其中木二糖和木三糖的含量高达94.7%。此外，从纯化后酶解低聚木糖产物显示可有效清除自由基，抗氧化活性>90%。综上所述，本研究通过木聚糖酶和阿拉伯呋喃糖苷酶Abf62A的协同生物催化作用，以绿色可持续的方式从麦秆木聚糖中制备低聚木糖，为人类和动物健康提供了一种益生元寡糖。

关键词: 阿拉伯呋喃糖苷酶, 木聚糖酶, 低聚木糖, 木聚糖, 生物合成

Abstract:

Xylo-oligosaccharides (XOSs) are a class of functional oligosaccharides whose main constituent is 2-7 xylose linked by β-1,4 glycosidic bonds, which are recognized as soluble dietary fibers with prebiotic activity. Recently, there is great interest to manufacture XOSs from xylan extracted from lignocellulosic biomass through the enzyme catalysis under the mild condition. In this work, the arabinofuranosidase Abf62A gene was cloned from Aspergillus usamii genomic DNA through sequential molecular processes and expressed in Pichia pastoris X33. The xylan (100 g/L) extracted xylan in wheat straw (WS) was biologically hydrolyzed into 50.32 g/L of XOSs by xylanase Xyn11A (200 U/g substrate) and arabinofuranase Abf62A (20 U/g substrate), which indicated a notable synergistic effect compared to the 34.44 g/L XOSs produced via Xyn11A. The 50.32 g/L of XOSs products comprised xylobiose (31.71 g/L), xylotriose (15.92 g/L), xylotetraose (1.65 g/L) and xylopentaose (1.04 g/L). Notably, the combined content of xylobiose and xylotriose accounted for up to 94.7%. The XOSs purified from the enzyme hydrolysate could effectually scavenge free radicals, and the antioxidant activity was more than 90%. To sum up, XOSs were biologically manufactured from wheat straw xylan through the synergistic biocatalysis via xylanase and arabinofuranosidase Abf62A in a green and sustainable way, rending one kind of prebiotic oligosaccharides with substantial positive effects on human and animal health.

Key words: arabinofuranosidase, xylanase, xylo-oligosaccharides, xylan, biosynthesis

中图分类号:

Q814

胡蝶, 徐道铸, 鲁志毅, 唐卫, 樊博, 何玉财. 木聚糖酶Xyn11A与阿拉伯呋喃糖苷酶Abf62A协同水解麦秆木聚糖生物合成低聚木糖[J]. 合成生物学, DOI: 10.12211/2096-8280.2025-037.

HU Die, XU Daozhu, LU Zhiyi, TANG Wei, FAN Bo, HE Yucai. Biosynthesis of xylo-oligosaccharides from wheat straw xylan via the synergistic hydrolysis by xylanase Xyn11A and arabinofuranosidase Abf62A[J]. Synthetic Biology Journal, DOI: 10.12211/2096-8280.2025-037.

图/表 9

Fig. 1 Cloning and expression analysis of Abf62A in P. pastoris

Fig. 2 Abf62A enzyme activity of different strains

Fig. 3 Effects of culture time (a) and temperature (b) on the enzyme activity of Abf62A fermentation liquor.

Fig. 4 SDS-PAGE results Abf62A protein(Lane A: Abf62A fermentation liquor; Lane B: Abf62A flow-through solution; Lane C: Abf62A eluate solution; Lane D: Abf62A pure enzyme solution)

Fig. 5 The optimum pH and pH stability of Abf62A (a) and its optimum temperature and temperature stability (b)

Fig. 6 Integrated process for valorizing wheat straw into antioxidant xylooligosaccharides

Table 1 Analysis of Xyn11A enzymatic hydrolysis products of different WS xylan

Product	UTX		PTX
Product	Concentration (g/L)	Proportion (%)	Concentration (g/L)	Proportion (%)
Xylose	0.63	3.08	1.37	3.97
Xylobiose	11.69	57.16	20.85	60.45
Xylotriose	7.61	37.21	11.44	33.17
Xylotetraose	0.41	2.01	1.13	3.28
Xylopentaose	0.26	1.27	1.0	2.90
Arabinose	0.17	0.83	0.43	1.25
Glucose	0.35	1.71	0.4	1.16

Table 2 Sugar content and proportion in xylan hydrolysates of different WS.

Product	UTX		PTX
Product	Concentration (g/L)	Proportion (%)	Concentration (g/L)	Proportion (%)
Xylose	0.55	2.35	1.43	2.63
Xylobiose	12.38	52.79	31.71	58.22
Xylotriose	8.35	35.61	15.92	29.23
Xylotetraose	0.62	2.64	1.65	3.03
Xylopentaose	0.47	2.01	1.04	1.91
Arabinose	0.62	2.64	2.34	4.30
Glucose	0.46	1.96	0.38	0.70

Fig. 7 Radical scavenging activity of xylo-oligosaccharides (XOS)(Data represent mean values of triplicate measurements.)

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