Release of signal peptide fragments into the cytosol requires cleavage in the transmembrane region by a protease activity that is specifically blocked by a novel cysteine protease inhibitor.
Signal peptides of secretory and membrane proteins are generated by proteolytic processing of precursor proteins after insertion into the endoplasmic reticulum membrane. Liberated signal peptides can be further processed, and the resulting N-terminal fragments are released toward the cytosol, where they may interact with target proteins like calmodulin. We show here that the processing of signal peptides requires a protease activity distinct from signal peptidase. This activity is inhibited specifically with a newly developed cysteine protease inhibitor, 1, 3-di-(N-carboxybenzoyl-l-leucyl-l-leucyl)amino acetone ((Z-LL)(2) ketone). Inhibitor studies revealed that the final, (Z-LL)(2) ketone-sensitive cleavage event occurs within the hydrophobic transmembrane region of the signal peptide, thus promoting the release of an N-terminal fragment into the cytosol.
Institut für Biochemie, ETH-Zentrum, Universitätstrasse 16, CH-8092 Zürich, Switzerland.
A
A
Weihofen
M K
MK
Lemberg
H L
HL
Ploegh
M
M
Bogyo
B
B
Martoglio
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D001667:Binding, Competitive; D002118:Calcium / Q000378:metabolism; D002147:Calmodulin / Q000378:metabolism; D002460:Cell Line; D003374:Coumarins / Q000494:pharmacology; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology*; D003600:Cytosol / Q000378:metabolism*; D004151:Dipeptides / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology*; D004591:Electrophoresis, Polyacrylamide Gel; D010450:Endopeptidases / Q000378:metabolism*; D004721:Endoplasmic Reticulum / Q000378:metabolism; D006801:Humans; D020128:Inhibitory Concentration 50; D007425:Intracellular Membranes / Q000378:metabolism; D049934:Isocoumarins; D007659:Ketones / Q000494:pharmacology*; D051379:Mice; D008969:Molecular Sequence Data; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D014176:Protein Biosynthesis; D021382:Protein Sorting Signals / Q000187:drug effects* / Q000502:physiology*; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D011691:Puromycin / Q000494:pharmacology; D015842:Serine Proteinase Inhibitors / Q000494:pharmacology; D013379:Substrate Specificity; D013997:Time Factors; D014158:Transcription, Genetic
C415363:1, 3-di-(N-carboxybenzoyl-leucyl-leucyl)amino acetone; D002147:Calmodulin; D003374:Coumarins; D015853:Cysteine Proteinase Inhibitors; D004151:Dipeptides; D049934:Isocoumarins; D007659:Ketones; D021382:Protein Sorting Signals; D011500:Protein Synthesis Inhibitors; D015842:Serine Proteinase Inhibitors; D011691:Puromycin; C046235:3,4-dichloroisocoumarin; D010450:Endopeptidases; D002118:Calcium