Testing an alternative expression system for protozoan proteins

ROMINA PETRIGH; ROSALIA MORETTA; PAULA RUYBAL; PABLO NUÑEZ; SILVINA WILKOWSKY; MARISA FARBER

Rosario

Otro; XLII Reunión anual de la Sociedad Argentina de Investigación en Bioquímica y Biología molecular; 2006

Sociedad Argentina de Investigación en Bioquímica y Biología molecular (SAIB)

Crithidia fasciculata is a non-pathogenic parasite of insects, which can be cultivated in high yields using inexpensive undefined serum-free media. The evolutionary proximity of C. fasciculata to protozoan pathogens, makes it an ideal vehicle for the expression of genes from lower eukaryotes. In this work we used C. fasciculata pNUS-H1 expression vectors (E. Tataud et al., 2002. Mol. Biochem. Parasitol. 120:196-204) for expressing the msa 2c gene from Babesia bovis, the causative agent of bovine babesiosis. The obtained constructs bore alternatively a poly-hystidine tag or a secretion peptide signal. To improve transfection efficiency different electroporationConditions were evaluated with GFP constructs.Transfected parasites were selected with Hygromycin B and resistant clones were obtained after 3 weeks. Up to now, stablelines were obtained and expression of MSA 2c was detected by Western blot. The expression of genes in environments that allow the production of functional proteins constitutes a main goal in order to improve the current expression tools for obtaining recombinant proteins. Concerning MSA 2c, the post- translational modifications should improve the antigenic efficacy in comparison with the same protein expressed in prokaryotic systems.