Cerda Liliana Alexandra, Valenzuela Susana Valeria, Diaz Pilar, Pastor Francisco I Javier
Department of Microbiology, Faculty of Biology, University of Barcelona, Barcelona, Spain.
Biotechnol Appl Biochem. 2016 Jan-Feb;63(1):51-6. doi: 10.1002/bab.1348. Epub 2015 Apr 28.
The gene coding for a lichenase from Paenibacillus barcinonensis BP-23, a powerful carbohydrate-degrading strain, was obtained using a genome walking strategy and expressed in Escherichia coli for further characterization. The amino acid sequence deduced from lic16A revealed that the lichenase is a single-domain enzyme belonging to the GH16 family. Purified recombinant Lic16A showed exclusive activity on β-1,3-1,4-glucans, showing a Km of 16.88 mg/mL and a Vmax of 266.09 U/mg using lichenan as a substrate. Lic16A was stable at 55 °C for at least 3 H in moderate pH conditions. Thin-layer chromatography analysis showed that the enzyme released a complex mixture of hydrolysis products, which consisted of different length oligosaccharides of intermediate mobility among cellooligomers. The health benefits of β-glucans's consumption and the increased interest for the use of their oligomers as prebiotics add interest to the study of Lic16A for the production of β-glucan-derived oligosaccharides and the evaluation of their biotechnological potential. This is the first report on β-1,3-1,4-glucanase produced by P. barcinonensis.
利用基因组步移策略获得了来自强大的碳水化合物降解菌株巴氏芽孢杆菌BP-23的地衣酶编码基因，并在大肠杆菌中表达以进行进一步表征。从lic16A推导的氨基酸序列表明，该酶是属于GH16家族的单结构域酶。纯化的重组Lic16A对β-1,3-1,4-葡聚糖具有专一活性，以地衣多糖为底物时，Km为16.88 mg/mL，Vmax为266.09 U/mg。Lic16A在中等pH条件下于55℃稳定至少3小时。薄层色谱分析表明，该酶释放出一种水解产物的复杂混合物，其由不同长度的低聚糖组成，这些低聚糖在纤维寡糖中具有中等迁移率。食用β-葡聚糖对健康有益，并且对将其低聚物用作益生元的兴趣增加，这使得对Lic16A用于生产β-葡聚糖衍生的低聚糖及其生物技术潜力评估的研究更具吸引力。这是关于巴氏芽孢杆菌产生的β-1,3-1,4-葡聚糖酶的首次报道。
