Heterologous Expression and Enzymatic Properties of αAgarase OUC-GaJJ96(α-琼胶酶OUC-GaJJ96的异源表达及酶学性质)
摘要
agarase plays an important role in the enzymatic preparation of functional agaro-oligosaccharides. However, there are only few studies focused on α-agarases. An α-agarase named OUC-GaJJ96 from Gilvimarinus agarilyticus was cloned, and its enzymatic properties together with hydrolysis products were studied. OUC-GaJJ96 belong to glycoside hydrolase 96 family with a molecular weight of approximately 180 kDa. The gene OUC-GaJJ96 comprised a 3759bp, encoding a protein of 1252 amino acids, with a signal peptide of 26 amino acids at its N-terminal. The enzymatic properties of OUC-GaJJ96 showed that the optimal temperature and pH were 30℃ and 7.0 (Tris-HCl buffer), respectively. The specific activity of OUC-GaJJ96 was 2.68U/mg. The analysis of HPLC and ESI-MS demonstrated that the hydrolysis products of OUC-GaJJ96 were agarobiose, agarotetraose and agarohexaose, in which agarotetraose was the main product. Therefore, OUC-GaJJ96 has potential for the manufacture of bioactive agaro-oligosaccharides.(琼胶酶在功能性琼胶寡糖的酶法制备中发挥着重要作用,目前有关α-琼胶酶的研究较少。克隆表达了来自溶藻性吉尔维菌(Gilvimarinus agarilyticus)的α-琼胶酶,命名为OUC-GaJJ96,并对其酶学性质及水解产物进行研究。该酶属于糖苷水解酶96家族,基因全长为3759bp,编码1252个氨基酸,N端含有26个氨基酸编码的信号肽,蛋白分子质量约为180kDa。酶学性质研究显示:OUC-GaJJ96最适反应温度和最适反应pH值分别为30℃和7.0(Tris-HCl缓冲液),比酶活力为2.68U/mg。通过高效液相色谱和电喷雾离子源质谱对OUC-GaJJ96的水解产物进行分析,结果表明:该酶水解产物是琼二糖、琼四糖和琼六糖,其中琼四糖是主要产物,OUC-GaJJ96可用于制备具有生物活性的琼胶寡糖。)