Nakamura N, Watanabe K, Horikoshi K
Biochim Biophys Acta. 1975 Jul 27;397(1):188-93. doi: 10.1016/0005-2744(75)90192-8.
Pullulanase (pullulan 6-glucanohydrolase EC 3.2.1.41) was purified about 290-fold from the culture fluid of Bacillus No. 202-1 by DEAE-cellulose adsorption, acetone fractionation, (NH4) 2SO4 precipitation and DEAE--cellulose column chromatography followed by Sephadex G-200 molecular sieve chromatography. The enzyme gave a single band of protein by disc polyacrylamide gel electrophoresis. The molecular weight was estimated as 92 000 by sodium dodecyl sulfate gel electrophoresis. The isolectric point was lower than pH 2.5. The optimum pH for enzyme action was about 8.5-9.0. The action of the enzyme on amylopectin and glycogen resulted in increase in the iodine coloration of 85% and 70%, respectively. The enzyme completely hydrolyzed 1,6-alpha-glucosidic linkages in amylopectin, glycogen and pullulan.
采用DEAE - 纤维素吸附、丙酮分级分离、硫酸铵沉淀、DEAE - 纤维素柱层析，随后进行Sephadex G - 200分子筛层析，从芽孢杆菌202 - 1号菌株的培养液中纯化出支链淀粉酶（支链淀粉6 - 葡聚糖水解酶，EC 3.2.1.41），纯化倍数约为290倍。该酶经圆盘聚丙烯酰胺凝胶电泳呈现单一蛋白条带。通过十二烷基硫酸钠凝胶电泳估计其分子量为92000。其等电点低于pH 2.5。该酶作用的最适pH约为8.5 - 9.0。该酶作用于支链淀粉和糖原后，碘染色分别增加了85%和70%。该酶能完全水解支链淀粉、糖原和支链淀粉中的1,6 - α - 糖苷键。
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