8-Nitroguanosine 3′,5′-cyclic monophosphate (8-nitro-cGMP) is the second messenger in nitric oxide/reactive oxygen species redox signaling. This molecule covalently binds to protein thiol groups, called S-guanylation, and exerts various biological functions. Recently, we have identified synaptosomal-associated protein 25 (SNAP-25) as a target of S-guanylation, and demonstrated that S-guanylation of SNAP25 enhanced SNARE complex formation. In this study, we have examined the effects of S-guanylation of SNAP-25 on the interaction between the SNARE complex and complexin (cplx), which binds to the SNARE complex with a high affinity. Pull-down assays and coimmunoprecipitation experiments have revealed that S-guanylation of Cys90 in SNAP-25 attenuates the interaction between the SNARE complex and cplx. In addition, blue native-PAGE followed by Western blot analysis revealed that the amount of cplx detected at a high molecular weight decreased upon 8-nitro-cGMP treatment in SH-SY5Y cells. These results demonstrated for the first time that S-guanylation of SNAP-25 attenuates the interaction between the SNARE complex and cplx.

中文翻译：

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8-Nitro-cGMP通过SNAP-25的S-Guanylation减弱SNARE复合物和Complexin之间的相互作用。

8-硝基鸟苷3'，5'-环一磷酸酯（8-nitro-cGMP）是一氧化氮/活性氧物种氧化还原信号传导的第二信使。该分子与蛋白硫醇基团共价结合，称为S-鸟苷酸化，并发挥各种生物学功能。最近，我们已经确定了突触体相关蛋白25（SNAP-25）作为S-胍基化的目标，并证明SNAP25的S-胍基化增强了SNARE复合物的形成。在这项研究中，我们已经研究了SNAP-25的S-鸟苷酸化对SNARE复合物和complexin（cplx）之间相互作用的影响，后者以很高的亲和力与SNARE复合物结合。下拉测定法和免疫共沉淀实验表明，SNAP-25中Cys90的S-鸟苷化减弱了SNARE复合物和cplx之间的相互作用。此外，蓝本色-PAGE，然后进行蛋白质印迹分析，结果表明，在SH-SY5Y细胞中进行8-硝基-cGMP处理后，高分子量检测到的cplx量减少。这些结果首次证明SNAP-25的S-鸟苷化减弱了SNARE复合物和cplx之间的相互作用。