Influence of the amino acid residue downstream of (Asp)(4)Lys on enterokinase cleavage of a fusion protein

We have studied the cleavage efficiency of the protease enterokinase (EK) u sing the novel vector pESP4. pESP4 is a yeast expression vector equipped wi th ligation-independent cloning sites, a GST purification tag, and a FLAG e pitope tag. EH is used to cleave the FLAG and GST tags leaving the protein of interest without any extraneously added amino acids. We have found that EK is relatively permissive of the amino acid residue downstream of the rec ognition sequence (the P-1' position). This makes EK an ideal choice to use as a protease to cleave any protein of interest cloned within the pESP4 ye ast expression vector. (C) 1999 Academic Press.