title: Prosomatostatin is proteolytically processed at the amino terminal segment by subtilase SKI-1.
abstract: Processing of prohormones to generate active products typically occurs at basic residues via cleavage by proprotein convertases. A less common type of cleavage is mediated at hydrophobic (L, V, F, N) or small amino acid (A, T, S) residues. Efforts to identify the proteinases responsible for processing precursors at their hydrophobic amino acids has led to the recent cloning of a new type-1 membrane-bound subtilase called SKI-1. The NH2-terminal region of prosomatostatin, previously shown to contain a sorting signal for the regulated secretory pathways, is processed to generate PSST[1-10]. The exact cleavage mechanism is unknown, but has been assumed to involve monobasic processing at Lys13 followed by carboxypeptidase trimming. We found that K13A mutation did not block PSST[1-10] production. Since the prosomatostatin sequence R8-Q9-F10-L11 \ qualifies as a potential SKI-1 substrate, using a vaccinia virus expression system along with HPLC and radioimmunoassays, we observed that overexpression of recombinant SKI-1 in COS-1 and HEK-293 cells significantly increased the production of PSST[1-10]. Additionally, in CHO cells lacking SKI-1, there was a significant reduction in PSST[1-10] production which could be increased upon SKI-1 stimulation. Mutagenesis studies showed that efficient processing of PSST to PSST[1-10] required the RXRXXL motif. However, this NH2-terminal cleavage was not a prerequisite for the formation of SST-14 and SST-28.
authors.affiliation: Fraser Laboratories, Department of Medicine, Room M3-15, Royal Victoria Hospital, McGill University, 687 Pine Avenue West, Montreal, Quebec, Canada H3A 1A1.
authors.forename: R
authors.initials: R
authors.lastname: Mouchantaf
authors.forename: H L
authors.initials: HL
authors.lastname: Watt
authors.forename: T
authors.initials: T
authors.lastname: Sulea
authors.forename: N G
authors.initials: NG
authors.lastname: Seidah
authors.forename: H
authors.initials: H
authors.lastname: Alturaihi
authors.forename: Y C
authors.initials: YC
authors.lastname: Patel
authors.forename: U
authors.initials: U
authors.lastname: Kumar
mesh_terms: D020816:Amino Acid Motifs; D000818:Animals; D016466:CHO Cells; D019556:COS Cells; D002851:Chromatography, High Pressure Liquid; D006224:Cricetinae; D006801:Humans; D007668:Kidney / Q000378:metabolism; D016296:Mutagenesis; D009154:Mutation / Q000235:genetics; D043484:Proprotein Convertases / Q000494:pharmacology*; D011498:Protein Precursors / Q000235:genetics / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D017434:Protein Structure, Tertiary; D011863:Radioimmunoassay; D012697:Serine Endopeptidases / Q000494:pharmacology*; D013004:Somatostatin / Q000235:genetics / Q000378:metabolism*; D014616:Vaccinia virus / Q000235:genetics
chemical_list: D011498:Protein Precursors; D013004:Somatostatin; C028456:prosomatostatin; D043484:Proprotein Convertases; D012697:Serine Endopeptidases; C115575:membrane-bound transcription factor peptidase, site 1