Purification and characteristics of fucoidanase obtained from Dendryphiella arenaria TM94
Purification and characteristics of fucoidanase obtained from Dendryphiella arenaria TM94
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DOI：
10.1007/s10811-010-9588-5
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发表时间：
2011-04
影响因子：
3.3
通讯作者：
Qianqian Wu;M. Zhang;Ke Wu;B. Liu;Jingmin Cai;R. Pan
中科院分区：
文献类型：
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作者：
Qianqian Wu;M. Zhang;Ke Wu;B. Liu;Jingmin Cai;R. Pan
Dendryphiella arenariaTM94 是一种专性海洋真菌。纯化TM94固态发酵表达的岩藻多糖酶。用柠檬酸缓冲液提取发酵的固体培养基，用丙酮沉淀提取物并在Sephadex G-100色谱上分离。纯化酶的比岩藻依聚糖酶活性是粗酶的27倍。酶的回收率为17.69%。使用SDS-PAGE来鉴定岩藻依聚糖酶的纯度和分子量。 SDS-PAGE凝胶上出现一条单条带，表明已获得相对纯的岩藻依聚糖酶。岩藻多糖酶的分子量为 180 kDa，等电点约为 pH 4.4。纯化的岩藻依聚糖酶似乎在 pH 6.0 时具有最大酶活性。 KM，使用墨角藻底物岩藻依聚糖时，酶的最大速度为 6.56 mg·mL−1 和 6.55 mg·mL−1·min−1。该酶可以是一种内切岩藻依聚糖酶，其可以将高分子量岩藻依聚糖水解成低分子量岩藻依聚糖而不是岩藻糖。
Dendryphiella arenariaTM94 is an obligate marine fungus. Fucoidanase expressed by TM94 by solid state fermentation was purified. The fermented solid medium was extracted with citric acid buffer, and the extracts were precipitated by acetone and separated on Sephadex G-100 chromatography. The specific fucoidanase activity of purified enzyme was 27-fold than that of the crude enzyme. The recovery of the enzyme was 17.69%. SDS-PAGE was used to identify the purity and the molecular weight of the fucoidanase. A single band appeared on SDS-PAGE gel which suggested that relatively pure fucoidanase has been obtained. The molecular weight of fucoidanase is 180 kDa and the isoelectric point was about pH 4.4. The purified fucoidanase appeared to have the maximum enzymatic activity at pH 6.0. KMand the maximum velocity of the enzyme was 6.56 mg·mL−1and 6.55 mg·mL−1·min−1by using fucoidan fromFucus vesiculosusas substrate. The enzyme may be a type of endo-fucoidanase which could hydrolyze high molecular weight fucoidan to low molecular weight fucoidan rather than to fucose.