Leishmania mexicana amazonensis: development of a peptide tag useful for labeling and purifying biotinylated recombinant proteins.
A number of peptide tags are available to facilitate the characterization of recombinant proteins. We have tested the bacterial oxaloacetate decarboxylase biotinylation domain for its efficacy in tagging recombinant proteins in vivo in Leishmania. To achieve efficient biotinylation, Leishmania also had to be co-transformed with the gene for bacterial biotin protein ligase (birA gene product). The recombinant chimeric protein could be detected on blots probed with avidin-horseradish peroxidase and purified on immobilized monomeric avidin resins.
Detke
Siegfried
S
Department of Biochemistry and Molecular Biology, University of North Dakota, Grand Forks, ND 58203, USA. sdetke@medicine.nodak.edu
D000818:Animals; D001360:Avidin / Q000737:chemistry; D019921:Biotinylation; D015153:Blotting, Western; D019731:Carbon-Nitrogen Ligases / Q000235:genetics / Q000378:metabolism; D002262:Carboxy-Lyases / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D018274:Electroporation; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism; D006735:Horseradish Peroxidase / Q000737:chemistry; D007711:Klebsiella pneumoniae / Q000201:enzymology; D007894:Leishmania mexicana / Q000235:genetics / Q000378:metabolism*; D010957:Plasmids; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D014170:Transformation, Genetic
D029968:Escherichia coli Proteins; C110928:NupM1 protein, Leishmania mexicana; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; D014157:Transcription Factors; D001360:Avidin; D006735:Horseradish Peroxidase; D002262:Carboxy-Lyases; C021916:oxaloacetate decarboxylase; D019731:Carbon-Nitrogen Ligases; C058125:birA protein, E coli