A new signal peptide useful for secretion of heterologous proteins from yeast and its application for synthetis of hirudin

Abstract The BGL2 gene from Saccharomyces cerevisiae encodes a β-glucanase which is localized to the yeast cell wall. The ability of a 23-amino acid (aa) signal peptide derived from the BGL2 gene to direct a heterologous protein to the secretory pathway of yeast has been compared to that of the MFα1 -encoded signal peptide in a series of gene fusions. As a model protein, the leech anticoagulant, recombinant hirudin variant 2-Lys 47 (HIR) has been studied. From a multicopy plasmid chimaeric proteins were produced which carry the BGL2 signal peptide (or the artificial BGL2 pre-Val 7 variant) ( i ) in front of the MFα1 pro sequence (or modified versions of MFα1 pro), i.e., a prepro signal, or ( ii ) joined directly to the heterologous protein. Accumulation of active HIR in yeast culture supernatants was observed when the BGL2 (or the BGL2 pre-Val 7 ) signal peptide were used in combination with either of three versions of the MFα1 pro peptide: the authentic MFα1 pro, a partially deleted MFα1 pro-Δ22–61, or a pro bearing an aa change (MFα1 pro-Gly 22 . In each case the BGL2 signal peptide (or its variant) has proven equally productive to the corresponding MFα1 peptide. Four times more active HIR was detected in the culture supernatant when either signal peptide was fused directly to the recombinant protein, as compared to a prepro protein version. Correct signal peptide cleavage was obtained when HIR was produced as a BGL2 pre-Val 7 :: fusion protein.