Han Wenjun, Zhao Shuai, Liu Huihui, Wu Zhihong, Gu Qianqun, Li Yuezhong
Key Laboratory of Marine Drugs, Chinese Ministry of Education, School of Medicine and Pharmacy, Ocean University of China, Qingdao 266003, China.
Wei Sheng Wu Xue Bao. 2012 Jun 4;52(6):776-83.
To isolate and identify a versatile carbohydrate-degrading bacterium from marine environments, and characterize the extracellular agarase activity.
The I2 staining method was applied in the isolation of agarose-degrading bacteria from coastal sediments of the Jiaozhou bay nearby Qingdao city, China. The JZB09 strain was cultured in multiple media using various complex polysaccharides as the sole carbon source to test the carbohydrate utilizing abilities. The 16S rRNA gene was cloned, sequenced and analyzed to identify the taxonomic position of the strain. Crude extracellular proteins were prepared using (NH4)2SO4 precepitation method. The dialyzed enzyme extract was applied in further studies including activity testing, activity staining, and agarose degrading for oligosaccharides purifiction. Three purified oligosaccharides were individually analyzed using thin layer chromatograph (TLC) and MALDI-TOF MS method.
The agarolytic marine bacterium, Persicobacter sp. JZB09, could use multiple complex polysaccharides as the sole carbon source and grew well on agarose, cellulose and xylan. The extracellular enzyme extract exhibits efficient and extensive degradation activity on agarose with an activity of 77.2 U/mg proteins. The extracellular agarase system (EAS) in the crude extracellular enzymes contains at least two agarose depolymerases with molecular masses of approximately 45 kDa and 70 kDa, respectively. A series of degradation products from agarose by the EAS was purified and identified as neoagaro-oligosaccharides, among which neoagarotetraose was the major product of the crude enzymatic products, which suggests that beta-agarase is the major constituent of the JZB09 EAS.
The polysaccharide-degrading bacterium Persicobacter sp. JZB09 and its polysaccharide-degrading system is promising for the exploration of polysaccharide depolymerase resources including beta-agarases.
从海洋环境中分离并鉴定一种多功能碳水化合物降解细菌，并对其胞外琼脂酶活性进行表征。
采用I2染色法从中国青岛市附近胶州湾的海岸沉积物中分离琼脂降解细菌。将JZB09菌株在多种培养基中培养，使用各种复合多糖作为唯一碳源来测试碳水化合物利用能力。克隆、测序并分析16S rRNA基因以确定该菌株的分类地位。使用硫酸铵沉淀法制备粗胞外蛋白。透析后的酶提取物用于进一步研究，包括活性测试、活性染色以及琼脂降解以纯化寡糖。使用薄层色谱（TLC）和基质辅助激光解吸电离飞行时间质谱（MALDI-TOF MS）方法分别对三种纯化的寡糖进行分析。
琼脂分解海洋细菌Persicobacter sp. JZB09可以使用多种复合多糖作为唯一碳源，并且在琼脂糖、纤维素和木聚糖上生长良好。胞外酶提取物对琼脂糖表现出高效且广泛的降解活性，活性为77.2 U/mg蛋白质。粗胞外酶中的胞外琼脂酶系统（EAS）至少包含两种琼脂糖解聚酶，分子量分别约为45 kDa和70 kDa。EAS对琼脂糖的一系列降解产物被纯化并鉴定为新琼脂寡糖，其中新琼脂四糖是粗酶产物的主要产物，这表明β-琼脂酶是JZB09 EAS的主要成分。
多糖降解细菌Persicobacter sp. JZB09及其多糖降解系统在探索包括β-琼脂酶在内的多糖解聚酶资源方面具有广阔前景。
