Choline-binding domain as a novel affinity tag for purification of fusion proteins produced in Pichia pastoris

The choline‐binding domain (ChoBD) of the carboxy‐terminal region of the Streptococcus pneumoniae amidase LYTA (C‐LYTA) presents a strong affinity for tertiary amines. We report a method for single‐step purification of proteins expressed in the methylotrophic yeast Pichia pastoris based on the fusio...

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Veröffentlicht in:	Biotechnology and bioengineering 2001-07, Vol.74 (2), p.164-171
Hauptverfasser:	Caubín, J., Martín, H., Roa, A., Cosano, I., Pozuelo, M., de la Fuente, J. M., Sánchez-Puelles, J. M., Molina, M., Nombela, C.
Format:	Artikel
Sprache:	eng
Schlagworte:	affinity tag Base Sequence beta-Lactamases - genetics beta-Lactamases - isolation & purification beta-Lactamases - metabolism Binding Sites Biological and medical sciences Biotechnology C-LYTA choline Choline - metabolism Chromatography, DEAE-Cellulose - methods Fundamental and applied biological sciences. Psychology Genetic engineering Genetic technics heterologous expression Methods. Procedures. Technologies Miscellaneous Molecular Sequence Data N-Acetylmuramoyl-L-alanine Amidase - genetics N-Acetylmuramoyl-L-alanine Amidase - isolation & purification N-Acetylmuramoyl-L-alanine Amidase - metabolism Pichia - genetics Pichia - metabolism Pichia pastoris protein purification Recombinant Fusion Proteins - genetics Recombinant Fusion Proteins - isolation & purification Recombinant Fusion Proteins - metabolism Streptococcus pneumoniae Streptococcus pneumoniae - enzymology Streptococcus pneumoniae - genetics Synthetic digonucleotides and genes. Sequencing tertiary amines yeast
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Zusammenfassung:	The choline‐binding domain (ChoBD) of the carboxy‐terminal region of the Streptococcus pneumoniae amidase LYTA (C‐LYTA) presents a strong affinity for tertiary amines. We report a method for single‐step purification of proteins expressed in the methylotrophic yeast Pichia pastoris based on the fusion of C‐LYTA to the protein of interest. We show that C‐LYTA can be efficiently expressed and secreted in this host. Tagged proteins fused to this binding domain can be purified on inexpensive DEAE matrices. It therefore provides a useful system for the purification of recombinant proteins with high specificity suitable for industrial purposes. © 2001 John Wiley & Sons, Inc. Biotechnol Bioeng 74: 164–171, 2001.
ISSN:	0006-3592 1097-0290
DOI:	10.1002/bit.1106