Production of SlpA protein from Lactobacillus brevis in Lactococcus lactis using a sugar inducible system.

HOLLMANN, AXEL; SAVIELLO, MARINAO; MYSHOSHI, ANDERSO; DELFEDERICO, LUCRECIA; SEMORILE, LILIANA; ACEVEDO, VASCO

Gotenburgo, Suecia

Congreso; FEMS 2009. Third Congress of European Microbiologists. Microbes and Man-interdependence and future challenges.; 2009

FEMS

Lactococcus lactis is a lactic acid bacterium that is widely used in the dairy industry for production and preservation of fermented foods. Since 1990s, many studies concerning the potential use of L. lactis for production and secretion of recombinant proteins were developed. This bacterium does not produce endotoxins, plasmid-free strains do not produce the protease PrtP, and only one protein Usp45 is secreted in detectable quantities, a feature that facilitates the purification and analysis of a protein of interest. Objetive: To obtain recombinant clones of L. lactis NCDO2118 able to express and secrete S-layer protein (SlpA) from L. brevis ATCC 8287. Methods: The XIES expression system was used. This system employs the Usp45 signal peptide and the xylose inducible xylT promoter of the xylose permease gene. Expression and secretion of transgenic SlpA was evaluated by Western blot assays by using anti-slpA antibodies. To evaluate the repression ability of the system, L. lactis containing pXYSEC:SlpA was grown in presence of non-PTS transported sugar galactose and two PTS-transported sugars, xylose and glucose, considered as inducer and repressor of the XIES system, respectively. Results: Western blot assays demonstrated the expression and secretion of the transgenic SlpA in L lactis. Expression characterization showed that the best results were obtained in stationary grown phase.  Versatility studies showed that after glucose addition, repression is quite effective and rapidly established, besides that elimination of xylose by washing showed not necessary. The possibility of repressing the expression without needing to deplete the inductor is a new and interesting feature of the pXIES system. Conclusions: In this work we have obtained a transformed L. lactis able to express SlpA in a secreted way from L. brevis by XIES expression system, as first approach to obtain genetically modified S-layer protein from lactobacilli for oral vaccine delivery systems.