P64: Proteomic analysis and Physiological Characterization of OMV(Outer Membrane Vesicle) of Pseudomonas putida KT2440

Outer membrane vesicles (OMVs) have been found from various pathogenic gram-negative bacteria such as Escherichia coli, Pseudomonas aeruginosa and Acinetobacter baumannii. In this study, we isolated OMV from a representative soil bacterium, Pseudomonas putida KT2440, which has biodegradability to various aromatic compounds. P.putida KT2440 was obtained from ATCC. Bacterial cultivation was performed according to previous methods(Yun et al. 2011). OMVs of P. putida KT2440 were purified from the bacterial culture supernatants by using a modified method from Kwon et al. LC-MS/MS analysis was performed according to a previous method(Park et al. 2006). Proteomic analysis of OMV showed outer membrane proteins (OmpF, OmpW, et al), TonB dependent receptors, outer membrane ferric siderophore receptor, and 17 kDa surface antigen as major outer membrane proteins of OMV of P. putida KT2440. OMV of P. putida KT2440 showed pathological effect on the culture cells originated from human lung cell. Production and its protein components of OMV were dependable to the carbon sources of culture media. Production of OMV was significantly increased in rich medium (LB) compared with in minimal medium containing benzoate and succinate as sole carbon sources, respectively. However, porins (benzoate-specific porin, BenF-like porin, et al) and enzymes (catechol 1,2-dioxygenase, benzoate dioxygenase, et al) for benzoate degradation were uniquely found in OMV prepared from P. putida KT2440 cultured in benzoate. This result suggests that production and components of OMV of P. putida KT2440 were controlled by the environmental stimulus or nutrients.