Selective and sensitive monitoring of caspase-1 activity by a novel bioluminescent activity-based probe.
The role of caspase-1 in inflammation has been studied intensely over recent years. However, the research of caspase-1 has remained difficult mainly due to the lack of sensitive and selective tools to monitor not only its abundance but also its activity. Here we present a bioluminescent activity-based probe (ABP) for caspase-1, developed by the Reverse Design concept, where chemically optimized protease inhibitors are turned into selective substrate ABPs. The probe exhibits excellent selectivity for caspase-1 and ∼1000-fold increase in sensitivity compared to available fluorogenic peptidic caspase-1 substrates. Moreover, we have been able to monitor and quantify specific caspase-1 activity directly in cell lysates. The activity correlated well with processing of prointerleukin-1β and prointerleukin-18 in phorbol 12-myristate 13-acetate (PMA)-stimulated cells. A detectable caspase-1 activity was present also in nonstimulated cells, consistent with processing of constitutively expressed prointerleukin-18.
Kindermann
Maik
M
Sanofi-Aventis Deutschland GmbH, R&D Chemical and Analytical Sciences, Industriepark Park Hoechst, Frankfurt, Germany.
Roschitzki-Voser
Heidi
H
Caglic
Dejan
D
Repnik
Urska
U
Miniejew
Catherine
C
Mittl
Peer R E
PR
Kosec
Gregor
G
Grütter
Markus G
MG
Turk
Boris
B
Wendt
K Ulrich
KU
D001224:Aspartic Acid / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000737:chemistry; D019086:Bridged Bicyclo Compounds, Heterocyclic / Q000138:chemical synthesis / Q000737:chemistry*; D020170:Caspase 1 / Q000235:genetics / Q000378:metabolism*; D061945:Caspase Inhibitors; D045744:Cell Line, Tumor; D015853:Cysteine Proteinase Inhibitors / Q000138:chemical synthesis / Q000737:chemistry*; D005456:Fluorescent Dyes / Q000138:chemical synthesis / Q000737:chemistry*; D006801:Humans; D020382:Interleukin-18 / Q000378:metabolism; D053583:Interleukin-1beta / Q000378:metabolism; D008163:Luminescent Measurements / Q000379:methods*; D011498:Protein Precursors / Q000378:metabolism; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D013379:Substrate Specificity
D019086:Bridged Bicyclo Compounds, Heterocyclic; C555051:CM 269; D061945:Caspase Inhibitors; D015853:Cysteine Proteinase Inhibitors; D005456:Fluorescent Dyes; D020382:Interleukin-18; D053583:Interleukin-1beta; D011498:Protein Precursors; D011994:Recombinant Proteins; D001224:Aspartic Acid; D020170:Caspase 1