DescriptionThe vector pMAL-p5X is designed to produce maltose-binding protein (MBP) fusions, where the protein of interest can be cleaved from MBP with the specific protease Factor Xa (NEB #P8010).
MBP fusions made with this vector include an N-terminal signal sequence, so the fusion protein is directed to the periplasm. The MBP has been engineered for tighter binding to amylose resin.
A gene or open reading frame is inserted into a restriction site of the vector polylinker, in the same translational reading frame as the malE gene (encoding maltose-binding protein). The fusion protein thus produced can be purified by amylose affinity chromatography. The sequence coding for the four amino acids Ile-Glu-Gly-Arg is present just upstream of the XmnI site. This allows the protein of interest to be cleaved from maltose-binding protein with the specific protease Factor Xa. Fragments inserted in the XmnI site (cleaves GAAGG↓ATTTC) will produce a fusion protein that, after Factor Xa cleavage, contains no vector-derived residues on the protein of interest.
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Properties and Usage
Affinity TagMaltose-Binding Protein (MBP)
Research Use Only
- The buyer/user has a non-exclusive license to use the vector for Research Purposes Only. Commercial use of this vector requires a license from New England Biolabs, Inc.
U.S. Patent No. 5,643,758
- NEB 10-beta E. coli (High Efficiency) (NEB #C3019) is recommended for propagation and subcloning. NEB Express Competent E.coli (High Efficiency) (NEB #C2523) is recommended for expression using this vector.
- Guan, C., Li, P., Riggs, P.D. and Inouye, H. (1987). Gene. 67, 21-30.
- Maina, C.V., Riggs, P.D., Grandea, A.G.III, Slatko, B.E., Moran, L.S., Tagliamonte, J.A., McReynolds, L.A. and Guan, C. (1988). Gene. 74, 365-373.
- Nagai, K. and Thogersen, H.C. (1987). Methods Enzymology. 153, 461-481.
- Riggs, P.D. (1990). Expression and Purification of Maltose-Binding Protein Fusions. In F.M. Ausebel, R. Brent, R.E. Kingston, D.D. Moore, J.G. Seidman, J.A. Smith and K. Struhl (Ed.), Methods Enzymology. 16.6.1-16.6.12. New York: John Wiley & Sons, Inc.
- Yanisch-Perron, C., Vieira, J. and Messing, J. (1985). Gene. 33, 103-119.