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 convertas e. Here we investigated the specificity of furin using synthetic pepti des based on the N-terminal sequence of human proalbumin. The propepti de was rapidly cleaved from the normal ((- 6)RGVFRR(- 1)DAPIKSEAVW(+9) ) peptide but as expected, there was no cleavage of the proalbumin Lil le analogue with a -2 His (-2H). Surprisingly, the effect of this subs titution could not be corrected by introducing a -4 Arg (-4R-2H). In c ontrast, the peptide -4R-2A was an excellent substrate being cleaved f ive times faster than normal, indicaabting that His is not allowed as an P-2 residue. Replacement of the -4 Val by Glu supported the expecte d 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 con tribution to cleavage, its replacement by Ala decreased the cleavage r ate to 60% of normal. The Lys-Arg propeptide was almost as good a subs trate as the normal Arg-Arg peptide, but the introduction of a Lys at P'(1) totally abolished processing. The exclusion of P'(1) positive ch arges would be an important requirement for preventing aberrant cleava ge in the middle of tetrabasic sequences.