A rapid fluorometric assay for the proteolytic activity of SKI‐1/S1P based on the surface glycoprotein of the hemorrhagic fever Lassa virus
FEBS letters, 2002•Wiley Online Library
The subtilase subtilisin kexin isozyme‐1 (SKI‐1)/site 1 protease (S1P), has been implicated
in the processing of Lassa virus glycoprotein C (GP‐C) precursor into GP1 and GP2 that are
responsible for viral fusion with the host cell membrane. Here, we studied in vitro the kinetics
of this cleavage by hSKI‐1 using an intramolecularly quenched fluorogenic (IQF) peptide, Q‐
GPC251–263 [Abz‐251Asp‐Ile‐Tyr‐Ile‐Ser‐Arg‐Arg‐Leu‐Leu↓ Gly‐Thr‐Phe‐Thr263‐3‐
NitroTyr‐Ala‐CONH2], containing the identified site. The measured V max (app)/K m (app) …
in the processing of Lassa virus glycoprotein C (GP‐C) precursor into GP1 and GP2 that are
responsible for viral fusion with the host cell membrane. Here, we studied in vitro the kinetics
of this cleavage by hSKI‐1 using an intramolecularly quenched fluorogenic (IQF) peptide, Q‐
GPC251–263 [Abz‐251Asp‐Ile‐Tyr‐Ile‐Ser‐Arg‐Arg‐Leu‐Leu↓ Gly‐Thr‐Phe‐Thr263‐3‐
NitroTyr‐Ala‐CONH2], containing the identified site. The measured V max (app)/K m (app) …
The subtilase subtilisin kexin isozyme‐1 (SKI‐1)/site 1 protease (S1P), has been implicated in the processing of Lassa virus glycoprotein C (GP‐C) precursor into GP1 and GP2 that are responsible for viral fusion with the host cell membrane. Here, we studied in vitro the kinetics of this cleavage by hSKI‐1 using an intramolecularly quenched fluorogenic (IQF) peptide, Q‐GPC251–263 [Abz‐251Asp‐Ile‐Tyr‐Ile‐Ser‐Arg‐Arg‐Leu‐Leu↓Gly‐Thr‐Phe‐Thr263‐3‐NitroTyr‐Ala‐CONH2], containing the identified site. The measured V max (app)/K m (app) was compared to those for other IQF SKI‐substrates. Q‐GPC251–263 is cleaved 10‐fold more efficiently than the previously known best SKI‐substrate, Q‐hproSKI134–142. This study confirmed the role of SKI‐1 in GP‐C processing and provides a novel, rapid and efficient enzymatic assay of SKI‐1.
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