Burnett School of Biomedical Sciences, College of Medicine, University of Central Florida, Orlando, FL 32816, USA.
Department of Physics, College of Sciences, University of Central Florida, Orlando, FL 32816, USA.
Toxins (Basel). 2019 Aug 4;11(8):458. doi: 10.3390/toxins11080458.
Protein disulfide isomerase (PDI) is mainly located in the endoplasmic reticulum (ER) but is also secreted into the bloodstream where its oxidoreductase activity is involved with thrombus formation. Quercetin-3-rutinoside (Q3R) blocks this activity, but its inhibitory mechanism against PDI is not fully understood. Here, we examined the potential inhibitory effect of Q3R on another process that requires PDI: disassembly of the multimeric cholera toxin (CT). In the ER, PDI physically displaces the reduced CTA1 subunit from its non-covalent assembly in the CT holotoxin. This is followed by CTA1 dislocation from the ER to the cytosol where the toxin interacts with its G protein target for a cytopathic effect. Q3R blocked the conformational change in PDI that accompanies its binding to CTA1, which, in turn, prevented PDI from displacing CTA1 from its holotoxin and generated a toxin-resistant phenotype. Other steps of the CT intoxication process were not affected by Q3R, including PDI binding to CTA1 and CT reduction by PDI. Additional experiments with the B chain of ricin toxin found that Q3R could also disrupt PDI function through the loss of substrate binding. Q3R can thus inhibit PDI function through distinct mechanisms in a substrate-dependent manner.
蛋白质二硫键异构酶（PDI）主要位于内质网（ER）中，但也会分泌到血液中，其氧化还原酶活性与血栓形成有关。槲皮素-3-鼠李糖苷（Q3R）阻断了这种活性，但它对 PDI 的抑制机制尚不完全清楚。在这里，我们研究了 Q3R 对另一个需要 PDI 的过程的潜在抑制作用：霍乱毒素（CT）的多聚体的解聚。在 ER 中，PDI 将还原型 CTA1 亚基从 CT 全毒素的非共价组装中物理置换。随后，CTA1 从 ER 易位到细胞质，毒素在细胞质中与其 G 蛋白靶标相互作用以产生细胞病变效应。Q3R 阻断了 PDI 与其结合的 CTA1 伴随的构象变化，这反过来又阻止了 PDI 从其全毒素中置换 CTA1，并产生了对毒素的抗性表型。CT 中毒过程的其他步骤不受 Q3R 影响，包括 PDI 与 CTA1 的结合以及 PDI 对 CT 的还原。用蓖麻毒素的 B 链进行的进一步实验发现，Q3R 也可以通过失去底物结合来破坏 PDI 功能。因此，Q3R 可以通过依赖底物的方式以不同的机制抑制 PDI 功能。
