YEAST-ASPARTIC-PROTEASE-3 (YAP3) PREFERS SUBSTRATES WITH BASIC RESIDUES IN THE P-2, P-1 AND P-2' POSITIONS

The yeast aspartic protease Yap3 is localised to the secretory pathway and correctly cleaves pro-alpha-mating factor at its dibasic sites, W e determined the specificity of Yap3 for mono-, di- and multi-basic cl eavage sites in the context of 15 residue synthetic proalbumin peptide s. Yap3 cleaved after dibasic ArgArg and LysArg sites but not after mo nobasic Arg sites even when there was an additional arginine at -6 and /or -4. Yap3 did not cleave a tetra-arginine site and tribasic sites ( RRR and RRK) were poor substrates. Cleavage always occurred C-terminal to the last arginine in the di- or tri-basic sequence. The optimal cl eavage site sequence was RR down arrow DR and this substrate was cleav ed 8-9-fold faster than the normal RR down arrow DA sequence. In contr ast to Kex2, Yap3 did not remove the propeptide from normal proalbumin or a range of natural or recombinant proalbumin variants, However at pH 4.0 Yap3 slowly cleaved proalbumin and albumin between domains 2 an d 3.