CLEAVAGE OF PROALBUMIN PEPTIDES BY FURIN REVEALS UNEXPECTED RESTRICTIONS AT THE P-2 AND P'(1), SITES

Proalbumin is the principal substrate of the in situ hepatic convertase. Here we investigated the specificity of furin using synthetic peptides based on the N-terminal sequence of human proalbumin. The propeptide was rapidly cleaved from the normal ((- 6)RGVFRR(- 1)DAPIKSEAVW(+9)) peptide but as expected, there was no cleavage of the proalbumin Lille analogue with a -2 His (-2H). Surprisingly, the effect of this substitution could not be corrected by introducing a -4 Arg (-4R-2H). In contrast, the peptide -4R-2A was an excellent substrate being cleaved five times faster than normal, indicaabting that His is not allowed as an P-2 residue. Replacement of the -4 Val by Glu supported the expected importance of a positive charge at P-4 as the cleavage rate dropped to 10% of normal after this substitution. The -6 Arg makes a small contribution to cleavage, its replacement by Ala decreased the cleavage rate to 60% of normal. The Lys-Arg propeptide was almost as good a substrate as the normal Arg-Arg peptide, but the introduction of a Lys at P'(1) totally abolished processing. The exclusion of P'(1) positive charges would be an important requirement for preventing aberrant cleavage in the middle of tetrabasic sequences.