Monday, August 13, 2012
Columbia Hall, Terrace Level (Washington Hilton)
Visceral leishmaniasis is a disease responsible for major impacts on public health, mainly in tropical and subtropical regions. Despite all the advances in molecular biology and immunology of infection, there is no vaccine available for leishmaniasis. Thus, there is a great need for production of specific antigens for the development of vaccines and diagnostic kits against this disease. The objective of this study was to evaluate the kinetics of expression of the 648 antigen of Leishmania infantum chagasi by Escherichia coli. The strain of E. coli expressing the 648 antigen was cultivated in 2xTY and TB media at 37ºC in a rotary shaker (200 rpm). In order to investigate the IPTG induction effect on growth and antigen expression, the cultures were induced by the addition of a final concentration of 1mM IPTG when the optical density reached 0.5. Results showed that the highest biomass concentration obtained was 5.45 g.L-1 in the TB medium and 3.13 g.L-1 in the 2xTY medium. Protein induction by IPTG showed that the best results were obtained in the 2xTY medium (0.21 g.L-1) after 6 h of cultivation. In the TB medium the maximum concentration of antigen obtained was 0.021 g.L-1 after 5 hours. It could be observed that antigen production was associated with growth behavior, with a maximum specific protein yield of 0.06 g.g-1 and 0.005 g.g-1 for the 2xTY and TB media, respectively. Protein expression was confirmed by SDS-PAGE, showing the antigen as a protein band with a molecular mass of approximately 24 kDa.