The 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.
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.
