Expression of foreign proteins in Trypanosoma congolense.
An expression vector was constructed to express foreign genes in Trypanosoma congolense. The foreign gene and a neomycin phosphotransferase (NPT) gene are flanked by glutamate and alanine rich protein (GARP) gene processing signals and their expression is driven by a ribosomal RNA gene promoter. The plasmid is not maintained as an episome in T. congolense, but the NPT gene permits selection of cells in which the plasmid has integrated into the genome. We used this plasmid to express luciferase, green fluorescent protein and a surface protein of Trypanosoma brucei, glycine-proline-glutamate glutamate threonine procyclic acidic repetitive protein (GPEET PARP). The plasmid-derived GPEET PARP is expressed on the surface of procyclic T. congolense and comigrates on a polyacrylamide gel with native GPEET PARP from T. brucei procyclic cells. We also attempted to use the plasmid to overexpress a previously identified T. congolense cysteine protease. The plasmid-derived cysteine protease mRNA species occurs in the transfected cells, but we were unable to detect increased levels of protein or protease activity.
Downey
N
N
Molecular Biology Ph.D. Program, University of Iowa, Iowa City, IA 52242, USA.
Donelson
J E
JE
D000595:Amino Acid Sequence; D000818:Animals; D003546:Cysteine Endopeptidases / Q000096:biosynthesis / Q000235:genetics; D005822:Genetic Vectors; D005839:Gentamicins / Q000494:pharmacology; D019868:Kanamycin Kinase / Q000235:genetics; D008562:Membrane Glycoproteins / Q000235:genetics; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000096:biosynthesis / Q000235:genetics; D010957:Plasmids; D014176:Protein Biosynthesis; D011506:Proteins / Q000235:genetics; D015800:Protozoan Proteins / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis*; D011995:Recombination, Genetic; D017386:Sequence Homology, Amino Acid; D014348:Trypanosoma congolense / Q000235:genetics*
C406724:GARP protein, Trypanosoma; D005839:Gentamicins; D008562:Membrane Glycoproteins; D009842:Oligopeptides; D011506:Proteins; D015800:Protozoan Proteins; D011994:Recombinant Proteins; C010680:antibiotic G 418; D019868:Kanamycin Kinase; D003546:Cysteine Endopeptidases