The propeptide is required for in vivo formation of stable active yeast proteinase A and can function even when not covalently linked to the mature region.
The PEP4-encoded aspartate protease proteinase A from Saccharomyces cerevisiae is synthesized as a zymogen (Ammerer, G., Hunter, C. P., Rothman, J. H., Saari, G. C., Valls, L. A., and Stevens, T. H. (1986) Mol. Cell. Biol. 6, 2490-2499; Woolford, C. A., Daniels, L. B., Park, F. J., Jones, E. W., van Arsdell, J. N., and Innis, M. A. (1986) Mol. Cell. Biol. 6, 2500-2510). We constructed a mutant form, lacking the propeptide. This form, still containing the signal peptide, was translocated into the endoplasmic reticulum, but the mature region was subsequently completely degraded. When a plasmid encoding only the propeptide after the signal peptide was introduced into a strain producing the mature region, a subpopulation of mature region molecules was rescued from the degradation and gained activity. Increased activity was found when the mature region was co-produced with increased amounts of propeptide, whereas truncated propeptides, lacking residues at its C terminus, were less efficient in the interaction with the mature region. We propose that the mature region of proteinase A cannot fold into its stable active conformation in the absence of the propeptide and that the propeptide can promote folding of the mature region, even when the propeptide and the mature region are not covalently linked.
van den Hazel
H B
HB
Department of Yeast Genetics, Carlsberg Laboratory, Copenhagen-Valby, Denmark.
Kielland-Brandt
M C
MC
Winther
J R
JR
D000483:Alleles; D000595:Amino Acid Sequence; D016282:Aspartic Acid Endopeptidases / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D003001:Cloning, Molecular; D004792:Enzyme Precursors / Q000235:genetics / Q000378:metabolism*; D005656:Fungal Proteins / Q000096:biosynthesis*; D005800:Genes, Fungal; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009838:Oligodeoxyribonucleotides; D010957:Plasmids; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000378:metabolism; D015183:Restriction Mapping; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins
D004792:Enzyme Precursors; D005656:Fungal Proteins; D009838:Oligodeoxyribonucleotides; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; C022325:aspartic proteinase A; C499267:PEP4 protein, S cerevisiae; D016282:Aspartic Acid Endopeptidases