Identification of specific tethered inhibitors for caspase-5.
The development of highly selective small molecule inhibitors for individual caspases, a class of cysteine-dependent aspartate-specific proteases, has been challenging due to conservation of the active site. Previously, we discovered an allosteric site at the dimer interface of caspases-3, -7, and -1 using disulfide trapping. Here, we show this approach can generate selective tethered ligands and inhibitors for caspase-5, which is remarkable considering its high sequence similarity to caspase-1. Among the 62 hits of a screen of ∼15 000 thiol-containing fragments, a naphthyl-thiazole-containing molecule was identified that selectively inhibited and labeled the allosteric cysteine in the p10 subunit of caspase-5, but caused very little inhibition or labeling of caspase-1. Interestingly, some of allosteric tethered compounds to caspase-5 did not inhibit its enzymatic activity, suggesting that thiol-labeling itself is not sufficient to drive inhibition. These studies validate an allosteric site on caspase-5 and provide a useful starting point to develop selective compounds to probe the role of caspase-5 separate from caspase-1 in the innate immune response.
Departments of Pharmaceutical Chemistry and Cellular and Molecular Pharmacology, University of California at San Francisco, 1700 4th Street, San Francisco, CA 94143, USA.
Junjun
J
Gao
James A
JA
Wells
D000495:Allosteric Site; D000595:Amino Acid Sequence; D020170:Caspase 1 / Q000737:chemistry / Q000378:metabolism; D061945:Caspase Inhibitors; D020169:Caspases / Q000235:genetics / Q000378:metabolism; D015853:Cysteine Proteinase Inhibitors / Q000138:chemical synthesis / Q000737:chemistry* / Q000302:isolation & purification / Q000494:pharmacology*; D004220:Disulfides / Q000737:chemistry; D004789:Enzyme Activation / Q000187:drug effects; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment
D061945:Caspase Inhibitors; D015853:Cysteine Proteinase Inhibitors; D004220:Disulfides; D011994:Recombinant Proteins; C099217:CASP5 protein, human; D020169:Caspases; D020170:Caspase 1