PMID- 10644685
OWN - NLM
STAT- MEDLINE
DCOM- 20000229
LR  - 20190508
IS  - 0021-9258 (Print)
IS  - 0021-9258 (Linking)
VI  - 275
IP  - 4
DP  - 2000 Jan 28
TI  - Biosynthesis and enzymatic characterization of human SKI-1/S1P and the processing
      of its inhibitory prosegment.
PG  - 2349-58
AB  - Biochemical and enzymatic characterization of the novel human subtilase hSKI-1
      was carried out in various cell lines. Within the endoplasmic reticulum of LoVo
      cells, proSKI-1 is converted to SKI-1 by processing of its prosegment into 26-,
      24-, 14-, 10-, and 8-kDa products, some of which remain tightly associated with
      the enzyme. N-terminal sequencing and mass spectrometric analysis were used to
      map the cleavage sites of the most abundant fragments, which were confirmed by
      synthetic peptide processing. To characterize its in vitro enzymatic properties, 
      we generated a secreted form of SKI-1. Our data demonstrate that SKI-1 is a
      Ca(2+)-dependent proteinase exhibiting optimal cleavage at pH 6.5. We present
      evidence that SKI-1 processes peptides mimicking the cleavage sites of the SKI-1 
      prosegment, pro-brain-derived neurotrophic factor, and the sterol regulatory
      element-binding protein SREBP-2. Among the candidate peptides encompassing
      sections of the SKI-1 prosegment, the RSLK(137)- and RRLL(186)-containing
      peptides were best cleaved by this enzyme. Mutagenesis of the latter peptide
      allowed us to develop an efficiently processed SKI-1 substrate and to assess the 
      importance of several P and P' residues. Finally, we demonstrate that, in vitro, 
      recombinant prosegments of SKI-1 inhibit its activity with apparent inhibitor
      constants of 100-200 nM.
FAU - Toure, B B
AU  - Toure BB
AD  - Laboratory of Biochemical, Protein Engineering Network of Centres of Excellence, 
      Clinical Research Institute of Montreal, University of Montreal, Montreal, Quebec
      H2W 1R7.
FAU - Munzer, J S
AU  - Munzer JS
FAU - Basak, A
AU  - Basak A
FAU - Benjannet, S
AU  - Benjannet S
FAU - Rochemont, J
AU  - Rochemont J
FAU - Lazure, C
AU  - Lazure C
FAU - Chretien, M
AU  - Chretien M
FAU - Seidah, N G
AU  - Seidah NG
LA  - eng
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PL  - United States
TA  - J Biol Chem
JT  - The Journal of biological chemistry
JID - 2985121R
RN  - 0 (DNA Primers)
RN  - EC 3.4.21.- (Proprotein Convertases)
RN  - EC 3.4.21.- (Serine Endopeptidases)
RN  - EC 3.4.21.112 (membrane-bound transcription factor peptidase, site 1)
SB  - IM
MH  - Amino Acid Sequence
MH  - Base Sequence
MH  - Cell Line
MH  - DNA Primers
MH  - Humans
MH  - Molecular Sequence Data
MH  - Peptide Mapping
MH  - *Proprotein Convertases
MH  - *Protein Processing, Post-Translational
MH  - Serine Endopeptidases/*biosynthesis/chemistry/metabolism
MH  - Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
EDAT- 2000/01/25 09:00
MHDA- 2000/03/04 09:00
CRDT- 2000/01/25 09:00
PHST- 2000/01/25 09:00 [pubmed]
PHST- 2000/03/04 09:00 [medline]
PHST- 2000/01/25 09:00 [entrez]
AID - 10.1074/jbc.275.4.2349 [doi]
PST - ppublish
SO  - J Biol Chem. 2000 Jan 28;275(4):2349-58. doi: 10.1074/jbc.275.4.2349.