Akiba S, Yamamoto K, Kumagai H
Biological Science Laboratories, Kao Corporation, Ibaraki, Japan.
Biosci Biotechnol Biochem. 1995 Jun;59(6):1048-51. doi: 10.1271/bbb.59.1048.
Three different carbohydrate-depleted enzymes were prepared from an endo-beta-1,4-glucanase of Aspergillus niger IFO31125 by treatment with endo-beta-N-acetylglucosaminidase or alpha-mannosidase. They were purified by Concanavalin A-Sepharose affinity and DEAE ion-exchange column chromatographies. The molecular sizes of these enzymes had been decreased from 40 kDa containing 9.0% carbohydrate to 39, 38, and 37 kDa with carbohydrate at 4.5, 1.3, and 0.8% (wt/wt), respectively. The native and these carbohydrate-depleted enzymes were compared in their enzymatic properties, and it was found that they were identical in their catalytic activities and both thermal and pH stabilities. However, the 37-kDa enzyme was more susceptible to proteolysis by Savinase, proteinase K, and Pronase E. On the other hand, the specific protease trypsin showed no such effect on activity of all enzymes. These results suggested that the core structure of the asparagine-linked sugar chain, which consisted of three monosaccharide residues, contributed to the high stability of the endo-beta-1,4-glucanase against protease digestion.
通过用内切-β-N-乙酰葡糖胺酶或α-甘露糖苷酶处理黑曲霉IFO31125的内切-β-1,4-葡聚糖酶，制备了三种不同的去糖基化酶。它们通过伴刀豆球蛋白A-琼脂糖亲和色谱和DEAE离子交换柱色谱进行纯化。这些酶的分子大小已从含9.0%碳水化合物的40 kDa分别降至含4.5%、1.3%和0.8%（重量/重量）碳水化合物的39 kDa、38 kDa和37 kDa。对天然酶和这些去糖基化酶的酶学性质进行了比较，发现它们在催化活性以及热稳定性和pH稳定性方面是相同的。然而，37 kDa的酶更容易被枯草杆菌蛋白酶、蛋白酶K和链霉蛋白酶E水解。另一方面，特异性蛋白酶胰蛋白酶对所有酶的活性均无此影响。这些结果表明，由三个单糖残基组成的天冬酰胺连接糖链的核心结构有助于内切-β-1,4-葡聚糖酶对蛋白酶消化具有高稳定性。
