Demidyuk I V, Nosovskaya E A, Tsaplina I A, Karavaiko G I, Kostrov S V
Lomonosov Moscow State Academy of Fine Chemical Technology, Moscow, Russia.
Biochemistry (Mosc). 1997 Feb;62(2):171-5.
Enzyme catalyzing hydrolysis of a substrate of Glu,Asp-specific proteinases (Z-Glu-pNA) and cleaving bond Glu13-Ala14 in the oxidized insulin B chain was purified to homogeneity from the culture medium of Thermoactinomyces species using hydrophobic chromatography on phenyl-Sepharose CL 4B as the key purification step. The molecular weight of the proteinase is 23 kD. The enzyme is completely inhibited by diisopropyl fluorophosphate and is stable at pH 5-11. The pH optimum for the hydrolysis of azocasein as substrate is 8.5. The temperature optimum for proteolytic activity is 55 degrees C. The N-terminal sequence of the proteinase is: Ser-Val-Leu-Gly-Thr-Asp-Glu-Arg-Thr-Arg-Val-Thr-Asn-Thr-Thr-Thr-Tyr-Pro- Tyr- Trp-.
从嗜热放线菌属的培养基中，通过以苯基琼脂糖凝胶CL 4B进行疏水层析作为关键纯化步骤，纯化出了一种能催化水解Glu、Asp特异性蛋白酶（Z-Glu-pNA）的底物并切割氧化胰岛素B链中Glu13-Ala14键的酶，使其达到了均一性。该蛋白酶的分子量为23 kD。该酶被二异丙基氟磷酸完全抑制，在pH 5 - 11时稳定。以偶氮酪蛋白为底物水解的最适pH为8.5。蛋白水解活性的最适温度为55℃。该蛋白酶的N端序列为：Ser-Val-Leu-Gly-Thr-Asp-Glu-Arg-Thr-Arg-Val-Thr-Asn-Thr-Thr-Thr-Tyr-Pro-Tyr-Trp- 。
