EFFECTS OF TEMPERATURE AND CYCLOHEXIMIDE ON SECRETION OF CLONED INVERTASE FROM RECOMBINANT SACCHAROMYCES-CEREVISIAE

The effects of temperature on the kinetics and efficiency of secretion of cloned invertase were investigated in a recombinant yeast system. This system consisted of the baker's yeast Saccharomyces cerevisiae (S EY2102) transformed with the 2 mu-based plasmid pBR58 which contains t he entire SUC2 gene including the promoter, signal sequence, and struc tural gene. The recombinant yeast produces the naturally secreted yeas t enzyme invertase, In transition experiments done at temperatures ran ging from 25 degrees to 45 degrees C, the maximum invertase level and secretion rate exhibited maxima of 5.5 U/mL . OD and 4.6 U/mL . OD per hour, respectively, at 35 degrees C. Experiments involving the use of cycloheximide showed that it took approximately 15 min for secreted i nvertase to move through the secretion pathway, which held 0.4 U/mL . OD of specific activity. (C) 1995 John Wiley & Sons, Inc.