Positional and additive effects of basic amino acids on processing of precursor proteins within the constitutive secretory pathway.

We have recently shown that the Arg/Lys-X-Lys/Arg-Arg or Arg/Lys-X-X-X-Lys/Arg-Arg sequence serves as a signal for cleavage of precursor proteins within the constitutive secretory pathway, and this cleavage is catalyzed by furin, a mammalian homolog of the yeast Kex2 protease. In this study, we further examined sequence requirements for the constitutive precursor cleavage. Based on the data concerning cleavage efficiencies of various prorenin mutants with amino acid substitution(s) around the native cleavage site expressed in CHO cells, we revised the sequence rules that govern the constitutive cleavage as follows: (i) the Arg residue at position -1 is essential; (ii) in addition to the Arg at position -1, at least two out of the three basic residues at positions -2, -4, and -6 are required for efficient cleavage (the presence of all the three basic residues results in most efficient cleavage); (iii) at position +1, a hydrophobic aliphatic amino acid is not suitable.