Purification and characterization of ginsenoside Ra-hydrolyzing β-D-Xylosidase from Bifidobacterium breve K-110, a human intestinal anaerobic bacterium
Purification and characterization of ginsenoside Ra-hydrolyzing β-D-Xylosidase from Bifidobacterium breve K-110, a human intestinal anaerobic bacterium
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DOI：
10.1248/bpb.26.1170
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发表时间：
2003-08-01
影响因子：
2
通讯作者：
Kim, DH
中科院分区：
文献类型：
--
作者：
Shin, HY;Lee, JH;Kim, DH
β-D-木糖苷酶(EC 3.2.1.37)是从人参皂苷Ra代谢短双歧杆菌K-110中分离得到的。经硫酸铵沉淀、QAE-纤维素柱层析、丁基香料醇层析、羟基磷灰石层析和Q-Sepharose柱层析等层析，β-D-木糖苷酶得到明显的纯化，最终比活力为51.8mumol/min/mg。经SDS-PAGE和凝胶过滤分析，β-D-木糖苷酶的相对分子质量为49 kDa，由单一亚基组成。β-D-木糖苷酶的最适pH为5.0，最适温度为37℃。纯化后的酶可被PCMS有效抑制。β-D-木糖苷酶对对-硝基苯基-β-D-吡喃木苷的作用最大，其次是人参皂苷Ra1和人参皂苷Ra2。该酶能将木聚糖水解为木糖，但不作用于对-硝基苯基-β-吡喃葡萄糖苷、对-硝基苯基-β-半乳糖苷或对-硝基苯基-β-D-岩藻糖苷。这些发现表明，这是首次报道从厌氧双歧杆菌中分离纯化人参皂苷水解酶。
beta-D-Xylosidase (EC 3.2.1.37) has been purified from ginsenoside Ra-metabolizing Bifidobacterium breve K-110, which was isolated from human intestinal microflora. beta-D-Xylosidase was purified to apparent homogeneity by a combination of ammonium sulfate precipitation, QAE-cellulose, butyl-toyopearl, hydroxyapatit and Q-Sepharose column chromatographies with the final specific activity of 51.8 mumol/min/mg. Molecular weight of beta- D-xylosidase is 49 kDa by SDS-PAGE and gel filtration, which consisted of a single subunit. beta-D-Xylosidase showed optimal activity at pH 5.0 and 37 degreesC. The purified enzyme was potently inhibited by PCMS. beta-D-Xylosidase acted to the greatest extent on p-nitrophenyl-beta-D-xylopyranoside, followed by ginsenoside Ra1 and ginsenoside Ra2. This enzyme hydrolyzed xylan to xylose, but did not act on p-nitrophenyl-beta-glucopyranoside, p-nitrophenyl-beta-galactopyranoside or p-nitrophenyl-beta-D-fucopyranoside. These findings suggest that this is the first reported purification of ginsenoside-hydrolyzing beta-D-Xylosidase from an anaerobic Bifidobacterium sp.