Study Of Quorum Sensing Gene In Klebsiella Pneumoniae

Quorum sensing (QS) represents a bacterial cell-cell communication mechanism by which bacteria coordinate their gene expression in a cell density-dependent manner. QS is mediated by small diffusible molecule termed autoinducer. Klebsiella pneumoniae is the most extensively studied among a number of 3-hydroxypropionic acid (3-HP) and 1,3-propanediol (1,3-PDO)-producing microorganisms. K. pneumoniae secrets type 2 signaling molecules.To characterize the function of QS gene in K. pneumoniae, the ygaG gene which encodes type 2 autoinducers (AI-2) synthase was PCR amplified and ligated to vector pET-pk. After transformation, a recombinant strain named K. pneumoniae(pET-ygaG) was generated. Meanwhile, the gene ygaG was deleted by RecA-dependent homologous recombination approach, resulting in a strain named K. pneumoniae △ygaG.The influence of ygaG deletion on protein expression in K. pneumoniae was investigated. Three differential protein bands at 28 kDa and 35 kDa were revealed by SDS-PAGE, MALDI-TOF-MS, and quantitative Real-time PCR. The results showed that the deletion of ygaG up-regulated or down-regulated the expression of genes related to membrane and metabolism. In other words, QS might influence membrane protein synthesis and metabolic pathways.We analyzed the biomass, glycerol consumption, and production of metabolites in a 24 h fermentation. It was showed that overexpression ofygaG inhibited K. pneumoniae growth, while knockout of ygaG benefited cell growth. Moreover, QS was found to affect metabolic flux, indicating that QS might control metabolic pathways in K. pneumoniae.Scanning electron microscope (SEM) was recruited to visualize the surface landscape of the mutant K. pneumoniae △ygaG. Compared to the wild type, the lines and wrinkles was shallower and outer membrane was smoother. On the other hand, through antimicrobial susceptibility test, the inhibition zones of the mutant were found to be smaller than that of wild type strain, suggesting that ygaG deletion strengthened the resistance of K. pneumoniae to drugs. Overall these findings suggest that QS behavior may be associated with biofilm formation and the pathogenicity of K. pneumoniae.