Expression of Bacillus subtilis neutral protease gene (nprE) in Saccharomyces cerevisiae

This study investigated the expression of the Bacillus subtilis neutral protease gene (nprE) in the yeast Saccharomyces cerevisiae. When the intact nprE gene was expressed using its native signal peptide, the protein accumulated intracellularly as an unprocessed precursor without secretion or processing. To enable secretion, researchers fused the yeast invertase signal peptide upstream of the mature NprE coding sequence. This hybrid construct successfully directed the protease into the yeast secretory pathway and the protein was secreted into culture medium. However, the secreted protein underwent extensive glycosylation and became biologically inactive. The findings demonstrate that bacterial signal peptides do not function effectively in yeast secretory pathways, and that post-translational glycosylation modifications in yeast can inactivate bacterial enzymes. The results suggest that while the yeast secretory system can process and secrete heterologous bacterial proteins when provided with appropriate targeting signals, the resulting modifications may render these proteins non-functional.

Key findings

• The native Bacillus subtilis signal peptide failed to direct nprE gene products into the yeast secretory pathway, resulting in intracellular accumulation of unprocessed pre-pro-NprE precursor
• Substituting the bacterial signal peptide with a yeast invertase signal peptide successfully achieved secretion of NprE protein into culture medium
• The secreted NprE protein underwent extensive N-linked glycosylation, making it biologically inactive despite successful secretion
• Post-translational glycosylation modifications in yeast differed from those on bacterial amylases, which retained activity in their glycosylated form