ABC转运蛋白介导嗜酸乳杆菌摄入、代谢水苏糖的分子机制研究

Stachyose is one of natural functional oligosaccharides which have been applied in the research and development of functional foods. Stachyose can be catabolized by Lactobacillus acidophilus as sole carbon source and promote their growth in the gastrointestinal tract of nutritional deficiency patients, and therefore benefit the health of the host. However, the efficient use of stachyose and probiotics-involved functional foods are hampered by the paucity of molecular mechanisms of stachyose uptake and catabolism by L.acidophilus. Therefore, the molecular mechanism on the uptake and catabolism of stachyose in L.acidophilus will be investigated in the proposed study, and the translocation and potential molecular mechanisms of ABC transporter mediated-stachyose uptake in L.acidophilus, which was related to periplasmic binding protein, transporter permease and transporter nucleotide binding protein, will be explored by employing modern molecular biology methods. New insights will be provided for the transport of other oligosaccharides across the membrane. Catabolic pathway of stachyose in L.acidophilus will be revealed in proteomic level, which may be benefit for the research and development of stachyose and probiotics-involved functional foods that targeted to improve intestinal health.

水苏糖是天然存在的功能性低聚糖，已在功能性食品研发中得到应用。在营养缺乏的肠道内，它可作为嗜酸乳杆菌生长的唯一碳源，通过摄入、代谢两个过程促其生长、增殖，达到有益宿主健康的目的。但水苏糖不能通过简单的扩散作用自由进入细胞，而嗜酸乳杆菌摄入、代谢水苏糖的作用机制仍不清楚，使得以水苏糖、益生菌为主要功能因子，靶向性改善肠道健康为目标的功能性食品的高效利用受到了影响。本项目旨在运用现代分子生物学手段，从周质结合、跨膜转运和胞内结构域功能三个方面，阐述ABC转运蛋白介导嗜酸乳杆菌摄入水苏糖的过程，所得结果对理解低聚糖类物质的跨膜转运具有重要的理论参考价值。利用比较蛋白质组学技术，从蛋白质水平，解析嗜酸乳杆菌代谢水苏糖的关键酶，进而揭示嗜酸乳杆菌中水苏糖的代谢途径，为改善肠道健康为靶标的功能性食品的研究与开发提供科学依据。

水苏糖能够调节肠道微生物群落的平衡，是一类具有改善人类健康功效的潜在益生元。水苏糖通过刺激肠道中嗜酸乳杆菌的生长，预防肠道损伤和调节免疫系统。然而，嗜酸乳杆菌代谢水苏糖的机制仍不清楚。本研究通过比较蛋白质组学分析了水苏糖促进益生菌生长的分子机制，发现16种水苏糖代谢相关的蛋白。结合MALDI TOF / TOF MS质谱鉴定了9个蛋白斑点，其中6个在水苏糖培养基中高度上调表达。这些上调的高丰度蛋白质被分类为PTS，ABC通道ECF转运蛋白，碳代谢，果糖和甘露糖代谢以及核苷酸切除修复。经过实时定量聚合酶链反应，酶学活性测定等试验完成转录水平和酶学活性的验证。同时，本研究提出了ABC通道ECF转运蛋白摄入水苏糖的新型结构模型，并系统比较了转运蛋白摄入维生素及水苏糖的异同点。此研究为进一步阐明水苏糖促进益生菌生长提供理论依据。