MULTICOPY OVEREXPRESSION OF BOVINE PANCREATIC TRYPSIN-INHIBITOR SATURATES THE PROTEIN-FOLDING AND SECRETORY CAPACITY OF SACCHAROMYCES-CEREVISIAE

Bovine pancreatic trypsin inhibitor (BPTI) was expressed and secreted from a synthetic gene as a model system for the study of protein folding and secretion in Saccharomyces cerevisiae. The efficiency of different leader sequences in directing BPTI secretion was examined, and up to 11 mu g/ml of active BPTI was secreted. In some fusion constructs, inefficient proteolytic processing by Kex2p, Ste13p, and signal peptidase were observed immediately adjacent to the BPTI N terminus. Insertion of dipeptide spacers improved endoproteolytic processing substantially but the level of secretion was unchanged, Overexpression from a 2-mu m multicopy vector results in essentially unchanged BPTI secretion as compared to expression from a single copy centromere vector. BPTI expressed from a multicopy vector accumulates intracellularly in an unfolded form, indicating that available secretory chaperones and foldases can be saturated by increasing the rate of BPTI synthesis. (C) 1995 Academic Press, Inc.