Ubiquitin-activating enzyme (E1), ubiquitin-conjugating enzyme (E2) and ubiquitin protein ligase (E3) are the key enzymes in protein ubiquitylation. PYR-41 is a pyrazone derivative that selectively inhibits E1 with an IC50 value less than 10 μM and a 95% inhibition was observed at the concentration of 50 μM. PYR-41 at 50 μM efficiently inhibited cyclin E degradation in vitro. PYR-41 reduced the level of E1∼Ub thioesters in retinal pigment epithelial cells with IC50 values ranging from 10-25 μM. The exposure of retinal pigment epithelial cells with PYR-41 (50 μM) for 30 min also blocked the accumulation of ubiquitin conjugates in response to the proteasome inhibitor ALLN. In U2OS cells, treatment of PYR-41 (50 μM) for 8 hours inhibited ubiquitylation and proteasomal degradation of a test substrate. In HeLa cells treated with EGF (100 ng/mL), pretreatment with PYR-41 for 15 min decreased the ligand-induced EGFR ubiquitylation. PYR-41 at 50 μM also reduced NF-κB activation in Hela cells by inhibiting the ubiquitylation of upstream signaling molecules and by blocking the proteasomal degradation of IκBα[3]. In a mouse model of cecal ligation and puncture (CLP)-induced sepsis, intravenous administration of PYR-41 (5 mg/kg) immediately after CLP significantly decreased serum levels of proinflammatory cytokines, including TNF-α, IL-1β, and IL-6, and the levels of organ injury markers, such as AST, ALT, and LDH. PYR-41 treatment also reduced myeloperoxidase activity and suppressed apoptosis and caspase-3 degradation in the lungs of septic mice as compared to DMSO-treated group. PYR-41 also significantly promoted 10-day survival in mice with sepsis from 42% to 83%[4].
作用机制
PYR-41 selectively inhibits E1 possibly by covalently modifying the active site cysteine of E1.
PYR-41 细胞研究
细胞系
浓度
检测类型
检测时间
活性说明
数据源
HI5 insect cells
Function assay
30 min
Inhibition of N-terminal GST-tagged human UAE-catalyzed UAE-Ub thioester intermediate formation expressed in HI5 insect cells after 30 mins by immunoblotting analysis, IC50=10 μM
23360215
HI5 insect cells
Function assay
Inhibition of N-terminal GST-tagged human UAE-catalyzed [32P]-AMP:[a-32P]-ATP exchange expressed in HI5 insect cells by TLC analysis, IC50=6.4 μM
23360215
insect cells
Function assay
Inhibition of mouse recombinant His6-tagged UAE-catalyzed UAE-Ub thioester intermediate formation expressed in insect cells by SDS-PAGE analysis, IC50=10 μM
Inhibition of N-terminal GST-tagged human UAE-catalyzed UAE-Ub thioester intermediate formation expressed in HI5 insect cells after 30 mins by immunoblotting analysis, IC50=10 μM
23360215
HI5 insect cells
Function assay
Inhibition of N-terminal GST-tagged human UAE-catalyzed [32P]-AMP:[a-32P]-ATP exchange expressed in HI5 insect cells by TLC analysis, IC50=6.4 μM
23360215
insect cells
Function assay
Inhibition of mouse recombinant His6-tagged UAE-catalyzed UAE-Ub thioester intermediate formation expressed in insect cells by SDS-PAGE analysis, IC50=10 μM
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