Nov. 5, 2018: 1:30 p.m. - 2 p.m.
The methylotrophic yeast, Pichia pastoris, has been genetically engineered to produce many heterologous proteins for industrial and research purposes. In order to secrete proteins for easier purification from the extracellular medium, the coding sequences of recombinant proteins are often fused to the Saccharomyces cerevisiae α-mating factor secretion signal. This α-mating factor secretion leader contains a 19 amino acid pre region followed by a 65 amino acid pro region. Using computer modeling of its predicted secondary structure as a guide, extensive site-directed mutagenesis of the 84 residue leader peptide was performed in order to determine the effects of various deletions and substitutions on the export of recombinant cargo proteins. Though some mutations clearly dampened protein expression, deletion of amino acids 57-70, corresponding to the last alpha helix of α-mating factor secretion signal, increased secretion of reporter proteins horseradish peroxidase and lipase at least 50%. These findings raise the possibility that the secretory efficiency of the leader can be further optimized in the future.