State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai 200237, China; College of Food Science and Engineering, Ocean University of China, Qingdao 266003, China.
College of Food Science and Engineering, Ocean University of China, Qingdao 266003, China.
Carbohydr Res. 2014 Mar 31;388:147-51. doi: 10.1016/j.carres.2014.02.019. Epub 2014 Feb 26.
AgWH50A, a novel β-agarase, was cloned from Agarivorans gilvus WH0801 by degenerate and nested PCR. It consists of 942 amino acids (105 kDa), including a 21-amino acid signal peptide. AgWH50A shares the highest amino acid sequence homology with AgaD02 from Agarivorans sp. QM38 (53%). The recombinant agarase gene was expressed in Escherichia coli and purified by affinity chromatography. Maximum enzymatic activity (Km 5.97 mg/mL and Vmax 0.781 U/mg) was observed at pH 6.0 and 30 °C. Using matrix-assisted laser desorption/ionisation-time-of-flight mass spectrometry, Fourier transform-nuclear magnetic resonance spectrometry and thin-layer chromatography, we analysed the hydrolysis products and concluded that AgWH50A is a neoagarotetraose-forming β-agarase, which can cleave agarose into neoagarotetraose. This novel agarase has potential applications in the industrial production of neoagarotetraose and provides a new agarose hydrolysis model for future research.
AgWH50A 是一种新型 β-琼脂酶，通过简并和嵌套 PCR 从 Agarivorans gilvus WH0801 中克隆得到。它由 942 个氨基酸（105 kDa）组成，包括一个 21 个氨基酸的信号肽。AgWH50A 与 Agarivorans sp. QM38 的 AgaD02 具有最高的氨基酸序列同源性（53%）。重组琼脂酶基因在大肠杆菌中表达，并通过亲和层析进行纯化。在 pH 值为 6.0 和 30°C 时，观察到最大酶活（Km 为 5.97 mg/mL，Vmax 为 0.781 U/mg）。通过基质辅助激光解吸/电离飞行时间质谱、傅里叶变换-核磁共振波谱和薄层色谱分析水解产物，我们得出结论，AgWH50A 是一种新型的 neoagarotetraose 形成 β-琼脂酶，能够将琼脂糖水解成 neoagarotetraose。这种新型的琼脂酶在 neoagarotetraose 的工业生产中具有潜在的应用价值，并为未来的研究提供了一种新的琼脂水解模型。
Zotero 插件