Teagasc, Moorepark Food Research Centre, Fermoy, County Cork, Ireland.
Appl Environ Microbiol. 2010 Jan;76(2):500-7. doi: 10.1128/AEM.01524-09. Epub 2009 Nov 20.
The exopolysaccharide beta-glucan has been reported to be associated with many health-promoting and prebiotic properties. The membrane-associated glycosyltransferase enzyme (encoded by the gtf gene), responsible for microbial beta-glucan production, catalyzes the conversion of sugar nucleotides into beta-glucan. In this study, the gtf gene from Pediococcus parvulus 2.6 was heterologously expressed in Lactobacillus paracasei NFBC 338. When grown in the presence of glucose (7%, wt/vol), the recombinant strain (pNZ44-GTF(+)) displayed a "ropy" phenotype, while scanning electron microscopy (SEM) revealed strands of polysaccharide-linking neighboring cells. Beta-glucan biosynthesis was confirmed by agglutination tests carried out with Streptococcus pneumoniae type 37-specific antibodies, which specifically detect glucan-producing cells. Further analysis showed a approximately 2-fold increase in viscosity in broth media for the beta-glucan-producing strain over 24 h compared to the control strain, which did not show any significant increase in viscosity. In addition, we analyzed the ability of beta-glucan-producing Lactobacillus paracasei NFBC 338 to survive both technological and gastrointestinal stresses. Heat stress assays revealed that production of the polysaccharide was associated with significantly increased protection during heat stress (60-fold), acid stress (20-fold), and simulated gastric juice stress (15-fold). Bile stress assays revealed a more modest but significant 5.5-fold increase in survival for the beta-glucan-producing strain compared to that of the control strain. These results suggest that production of a beta-glucan exopolysaccharide by strains destined for use as probiotics may afford them greater performance/protection during cultivation, processing, and ingestion. As such, expression of the gtf gene may prove to be a straightforward approach to improve strains that might otherwise prove sensitive in such applications.
β-葡聚糖多糖已被报道与许多促进健康和益生元特性相关。负责微生物β-葡聚糖产生的膜相关糖基转移酶（由 gtf 基因编码），催化糖核苷酸向β-葡聚糖的转化。在这项研究中，来自肠膜明串珠菌 2.6 的 gtf 基因在副干酪乳杆菌 NFBC338 中异源表达。当在葡萄糖（7%，wt/vol）存在下生长时，重组菌株（pNZ44-GTF(+））表现出“粘性”表型，而扫描电子显微镜（SEM）显示多糖链连接相邻细胞。通过与肺炎链球菌 37 型特异性抗体进行的凝集试验证实了β-葡聚糖的生物合成，该抗体特异性检测产葡聚糖的细胞。进一步分析表明，与对照菌株相比，产β-葡聚糖的副干酪乳杆菌 NFBC338 在 24 小时内培养基中的粘度增加了约 2 倍，而对照菌株的粘度没有明显增加。此外，我们分析了产β-葡聚糖的副干酪乳杆菌 NFBC338 对技术和胃肠道压力的生存能力。热应激试验表明，多糖的产生与热应激（60 倍）、酸应激（20 倍）和模拟胃液应激（15 倍）期间显著增加的保护作用相关。胆盐应激试验表明，与对照菌株相比，产β-葡聚糖的菌株的存活率提高了 5.5 倍。这些结果表明，益生菌候选菌株产生β-葡聚糖外多糖可能会在培养、加工和摄入过程中为它们提供更大的性能/保护。因此，gtf 基因的表达可能是一种简单的方法，可以改善在这些应用中可能敏感的菌株。
