Characterization of a novel and specific inhibitor for the pro-apoptotic protease Omi/HtrA2.
Omi/HtrA2 is a mammalian serine protease with high homology to bacterial HtrA chaperones. Omi/HtrA2 is localized in mitochondria and is released to the cytoplasm in response to apoptotic stimuli. Omi/HtrA2 induces cell death in a caspase-dependent manner by interacting with the inhibitor of apoptosis protein as well as in a caspase-independent manner that relies on its protease activity. We describe the identification and characterization of a novel compound as a specific inhibitor of the proteolytic activity of Omi/HtrA2. This compound (ucf-101) was isolated in a high throughput screening of a combinatorial library using bacterially made Omi-(134-458) protease and fluorescein-casein as a generic substrate. ucf-101 showed specific activity against Omi/HtrA2 and very little activity against various other serine proteases. This compound has a natural fluorescence that was used to monitor its ability to enter mammalian cells. ucf-101, when tested in caspase-9 (-/-) null fibroblasts, was found to inhibit Omi/HtrA2-induced cell death.
Cilenti
Lucia
L
Biomolecular Science Center and Department of Molecular Biology and Microbiology, University of Central Florida, Orlando, Florida 32826, USA.
Lee
Younghee
Y
Hess
Sibylle
S
Srinivasula
Srinivasa
S
Park
Kwon Moo
KM
Junqueira
Daniela
D
Davis
Hedvika
H
Bonventre
Joseph V
JV
Alnemri
Emad S
ES
Zervos
Antonis S
AS
D017209:Apoptosis; D004591:Electrophoresis, Polyacrylamide Gel; D016650:Fluorescein-5-isothiocyanate; D000074923:High-Temperature Requirement A Serine Peptidase 2; D006868:Hydrolysis; D024101:Mitochondrial Proteins; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D012697:Serine Endopeptidases / Q000187:drug effects* / Q000378:metabolism; D015842:Serine Proteinase Inhibitors / Q000494:pharmacology*
D024101:Mitochondrial Proteins; D011994:Recombinant Proteins; D015842:Serine Proteinase Inhibitors; D012697:Serine Endopeptidases; D000074923:High-Temperature Requirement A Serine Peptidase 2; D016650:Fluorescein-5-isothiocyanate