Proteolytic events in the processing of secreted proteins in fungi.
Secreted heterologous proteins have been found to be produced much less efficiently by fungi than secreted homologous ones. This could be due, at least in part, to proteolytic cleavage by site-specific endoproteases of the secretory pathway, similar to the yeast KEX2 protease and the mammalian dibasic endoproteinases found in secretory pathways. Mature secreted fungal proteins may be protected from such cleavage due to the absence of cleavable sites in exposed regions. A comparison of the dipeptide distributions of 33 secreted and 34 cytoplasmic proteins from fungal producers of extracellular enzymes indicated a significant bias for some doublets, including the basic dipeptides Lys-Arg, Arg-Arg and Arg-Lys which have also been demonstrated to be KEX2 substrates. Other combinations were also found to be rare in secreted proteins, which could indicate either a broader specificity of the considered endopeptidase, or the presence either in the secretory organelles or among the secreted proteins of additional proteases with different specificities. Experimental evidence that the Lys-Arg site is processed in Tolypocladium geodes was provided by cloning a synthetic prosequence upstream of a phleomycin resistance (Sh ble) gene and analyzing the N-terminus of the corresponding protein purified from the culture supernatant. This system also provides a tool for further studies of specific proteases of fungi.
Laboratoires CAYLA, Toulouse, France.
T P
TP
Calmels
F
F
Martin
H
H
Durand
G
G
Tiraby
D000595:Amino Acid Sequence; D001483:Base Sequence; D004352:Drug Resistance, Microbial; D010450:Endopeptidases / Q000378:metabolism*; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D005658:Fungi / Q000201:enzymology* / Q000235:genetics; D008969:Molecular Sequence Data; D010692:Phleomycins / Q000494:pharmacology; D011499:Protein Processing, Post-Translational; D021382:Protein Sorting Signals / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013379:Substrate Specificity
D005656:Fungal Proteins; D010692:Phleomycins; D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins; D010450:Endopeptidases