Expression and one-step purification of recombinant proteins using an alternative episomal vector for the expression of N-tagged heterologous proteins in Pichia pastoris.
Here we report the construction of an alternative episomal vector, pBGP3, which allows the expression of heterologous proteins with N-terminal hexahistidine and myc-epitope tags in Pichia pastoris. To test the usefulness of pBGP3, four cellulases from termites were expressed. Production was confirmed by activity assays and Western blot using anti-c-Myc antibody. Purification was performed by single-step Ni(2+)-affinity chromatography, which confirmed the efficiency of pBGP3.
Department of Biotechnology, The University of Tokyom, Tokyo, Japan.
Cristiane Akemi
CA
Uchima
Manabu
M
Arioka
D000818:Animals; D015153:Blotting, Western; D044602:Cellulases / Q000235:genetics; D002846:Chromatography, Affinity; D005822:Genetic Vectors / Q000235:genetics*; D006639:Histidine; D020049:Isoptera / Q000201:enzymology; D008722:Methods; D009842:Oligopeptides; D010843:Pichia / Q000235:genetics* / Q000378:metabolism; D010957:Plasmids; D016271:Proto-Oncogene Proteins c-myc; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000302:isolation & purification
C471213:His-His-His-His-His-His; D009842:Oligopeptides; D016271:Proto-Oncogene Proteins c-myc; D011994:Recombinant Proteins; D006639:Histidine; D044602:Cellulases