Kurakake M, Yo-u S, Nakagawa K, Sugihara M, Komaki T
Department of Food Science & Technology, Fukuyama University, Sanzou, Gakuenchou 1 banchi, Fukuyama, Hiroshima 729-0292, Japan.
Curr Microbiol. 2000 Jan;40(1):6-9. doi: 10.1007/s002849910002.
Chitosanase from Bacillus cereus S1 was purified, and the enzymatic properties were investigated. The molecular weight was estimated to 45,000 on SDS-PAGE. Optimum pH was about 6, and stable pH in the incubation at 40 degrees C for 60 min was 6-11. This chitosanase was stable in alkaline side. Optimum temperature was around 60 degrees C, and enzyme activity was relatively stable below 60 degrees C. The degradations of colloidal chitosan and carboxymethyl cellulose (CMC) were about 30 and 20% relative to the value of soluble chitosan, respectively, but colloidal chitin and crystalline cellulose were not almost hydrolyzed. On the other hand, S1 chitosanase adsorbed on colloidal chitin completely and by about 50% also on crystalline cellulose, in contrast to colloidal chitosan, which it did not adsorb. S1 chitosanase finally hydrolyzed 100% N-deacetylated chitosan (soluble state) to chitobiose (27.2%), chitotriose (40.6%), and chitotetraose (32.2%). In the hydrolysis of various chitooligosaccharides, chitobiose and chitotriose were not hydrolyzed, and chitotetraose was hydrolyzed to chitobiose. Chitobiose and chitotriose were released from chitopentaose and chitohexaose. From this specificity, it was hypothesized that the active site of S1 chitosanase recognized more than two glucosamine residues posited in both sides against splitting point for glucosamine polymer.
对蜡样芽孢杆菌S1的壳聚糖酶进行了纯化，并对其酶学性质进行了研究。在SDS-PAGE上估计其分子量为45,000。最适pH约为6，在40℃孵育60分钟时的稳定pH为6至11。这种壳聚糖酶在碱性方面稳定。最适温度约为60℃，酶活性在60℃以下相对稳定。相对于可溶性壳聚糖的值，胶体壳聚糖和羧甲基纤维素（CMC）的降解率分别约为30%和20%，但胶体几丁质和结晶纤维素几乎未被水解。另一方面，与不吸附的胶体壳聚糖相反，S1壳聚糖酶完全吸附在胶体几丁质上，对结晶纤维素的吸附率也约为50%。S1壳聚糖酶最终将100%的N-脱乙酰壳聚糖（可溶状态）水解为壳二糖（27.2%）、壳三糖（40.6%）和壳四糖（32.2%）。在各种壳寡糖的水解中，壳二糖和壳三糖未被水解，壳四糖被水解为壳二糖。壳二糖和壳三糖从壳五糖和壳六糖中释放出来。基于这种特异性，推测S1壳聚糖酶的活性位点识别在氨基葡萄糖聚合物的切割点两侧排列的两个以上的氨基葡萄糖残基。
