O. Schonberger et al., SPLIT INVERTASE POLYPEPTIDES FORM FUNCTIONAL COMPLEXES IN THE YEAST PERIPLASM IN-VIVO, Proceedings of the National Academy of Sciences of the United Statesof America, 93(18), 1996, pp. 9612-9617
The assembly of functional proteins from fragments in vivo has been re
cently described for several proteins, including the secreted maltose
binding protein in Escherichia coil. Here we demonstrate for the first
time that split gene products can function within the eukaryotic secr
etory system. Saccharomyces cerevisiae strains able to use sucrose pro
duce the enzyme invertase, which is targeted by a signal peptide to th
e central secretory pathway and the periplasmic space. Using this enzy
me as a model we find the following: (i) Polypeptide fragments of inve
rtase, each containing a signal peptide, are independently translocate
d into the endoplasmic reticulum (ER) are modified by glycosylation, a
nd travel the entire secretory pathway reaching the yeast periplasm. (
ii) Simultaneous expression of independently translated and translocat
ed overlapping fragments of invertase leads to the formation of an enz
ymatically active complex, whereas individually expressed fragments ex
hibit no activity. (iii) An active invertase complex is assembled in t
he ER, is targeted to the yeast periplasm, and is biologically functio
nal, as judged by its ability to facilitate growth on sucrose as a sin
gle carbon source. These observations are discussed in relation to pro
tein folding and assembly in the ER and to the trafficking of proteins
through the secretory pathway.