Defective Escherichia coli signal peptides function in yeast.
To investigate structural characteristics important for eukaryotic signal peptide function in vivo, a hybrid gene with interchangeable signal peptides was cloned into yeast. The hybrid gene encoded nine residues from the amino terminus of the major Escherichia coli lipoprotein, attached to the amino terminus of the entire mature E. coli beta-lactamase sequence. To this sequence were attached sequences encoding the nonmutant E. coli lipoprotein signal peptide, or lipoprotein signal peptide mutants lacking an amino-terminal cationic charge, with shortened hydrophobic core, with altered potential helicity, or with an altered signal-peptide cleavage site. These signal-peptide mutants exhibited altered processing and secretion in E. coli. Using the GAL10 promoter, production of all hybrid proteins was induced to constitute 4-5% of the total yeast protein. Hybrid proteins with mutant signal peptides that show altered processing and secretion in E. coli, were processed and translocated to a similar degree as the non-mutant hybrid protein in yeast (approximately 36% of the total hybrid protein). Both non-mutant and mutant signal peptides appeared to be removed at the same unique site between cysteine 21 and serine 22, one residue from the E. coli signal peptidase II processing site. The mature lipo-beta-lactamase was translocated across the cytoplasmic membrane into the yeast periplasm. Thus the protein secretion apparatus in yeast recognizes the lipoprotein signal sequence in vivo but displays a specificity towards altered signal sequences which differs from that of E. coli.
Pines
O
O
Department of Biochemistry, State University of New York, Stony Brook 11794.
Lunn
C A
CA
Inouye
M
M
D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics*; D005796:Genes; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D010957:Plasmids; D021382:Protein Sorting Signals / Q000235:genetics* / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D001618:beta-Lactamases / Q000235:genetics*
D021382:Protein Sorting Signals; D001618:beta-Lactamases