Localization of cloned invertase in Saccharomyces cerevisiae directed by the SUC2 and MFalpha1 signal sequences.
Protein localization in Saccharomyces cerevisiae was studied with two plasmid systems used as a model: one containing the SUC2 structural gene fused with the MFalpha1 (alpha-factor) promoter and signal-sequence, the other containing the entire SUC2 gene. Special emphasis was placed on the effect of promoter/signal-sequence (SUC2 vs. MFalpha1) on the efficiency of invertase transport. The MFalpha 1 and SUC2 signal sequences were capable of transporting, respectively, 83% and 77% of cloned invertase out of the cytoplasm. However, the SUC2 promoter was easier to control since a six-fold enhancement of the transported invertase activity associated with derepression was achieved in response to a glucose concentration change from 10 to 2 g/L Cloning on a multicopy plasmid resulted in a four-fold increase in total specific invertase activity over the wild type yeast strain (which harbors a single copy of the SUC2 gene on the chromosome), whereas the chromosomal site was more efficient for invertase localization yielding over 90% of the invertase transported out of the cytoplasm. Transient experiments done with the SUC2 signal-sequence-containing plasmid showed that the specific invertase activity in the periplasmic space reached a maximum three hours after derepression, then decreased very slowly with an accompanying gradual increase in invertase activity in the growth medium.
Marten
M R
MR
School of Chemical Engineering, Purdue University, West Lafayette, Indiana 47907, USA.
Seo
J H
JH