OVEREXPRESSION OF BINDING-PROTEIN AND DISRUPTION OF THE PMR1 GENE SYNERGISTICALLY STIMULATE SECRETION OF BOVINE PROCHYMOSIN BUT NOT PLANT THAUMATIN IN YEAST

When the heterologous proteins thaumatin and bovine prochymosin are pr oduced in yeast cells as a fusion with the yeast invertase secretory s ignal peptide, less than 2% of the product is secreted in a biological ly active form into the medium. The remainder accumulates intracellula rly in a misfolded conformation. We investigated whether this poor sec retion can be improved by overexpression of binding protein (BiP) one of the major chaperones in eukaryotic cells. Indeed, a tenfold increas e in the level of binding protein, as a result of the introduction of extra copies of the kar2 gene into yeast cells containing a single, in tegrated copy of the invertase/prochymosin fusion gene, caused more th an a 20-fold increase in the amount of extracellular prochymosin. By a dditional disruption of the PMR1 gene of these cells we were able to o btain secretion of virtually all of the prochymosin produced. Export o f thaumatin, on the other hand, was not significantly stimulated by bi nding protein overexpression.