Non-pathogenic trypanosomatid protozoa as a platform for protein research and production.

All currently existing eukaryotic protein expression systems are based on autonomous life forms. To exploit the potential practical benefits associated with parasitic organisms we have developed a new protein expression system based on Leishmania tarentolae (Trypanosomatidae), a protozoan parasite of lizards. To achieve strong transcription, the genes of interest were integrated into the small subunit ribosomal RNA gene. Expression levels obtained were up to 30 mg of recombinant protein per liter of suspension culture and increased linearly with the number of integrated gene copies. To assess the system's potential for production of post-translationally modified proteins, we have expressed human erythropoietin in L. tarentolae. The recombinant protein isolated from the culture supernatants was biologically active, natively processed at the N-terminus, and N-glycosylated. The N-glycosylation was exceptionally homogeneous, with a mammalian-type biantennary oligosaccharide and the Man(3)GlcNAc(2) core structure accounting for >90% of the glycans present. L. tarentolae is thus the first described biotechnologically useful unicellular eukaryotic organism producing biantennary fully galactosylated, core-alpha-1,6-fucosylated N-glycans.