Wnts是富含半胱氨酸的脂质修饰蛋白,调节发育过程,并参与各种病理状况。Wnts结构像一只手,具有棕榈酰基拇指和与卷曲(FZ)受体相互作用的食指状域。先前的研究表明棕榈酚基团和二硫化物在Wnt折叠中的重要性,分泌,和功能,但是结构基础还没有完全理解。这里,我们利用经典分子动力学模拟(总共800-ns)来研究拇指棕榈酰及其保守的二硫化物(183-190,181-195)如何调节Wnt-FZ相互作用和结构动力学。使用转向分子动力学实验,然后进行放松程序,我们还探索了这些二硫化物在Wnt-FZ复合物形成中是否重要。根据我们的结果,棕榈酰基团显著有助于稳定Wnt-FZ相互作用,和二硫化物调节这种贡献。我们还证明了二硫键183-190调节Wnt拇指波动,氢键网络,和二级结构。DCCM分析描绘了二硫键183-190在调节棕榈烯化环中的天然样集体运动中的作用,在去除二硫化物后发生了变化。拉伸松弛实验表明,两种二硫化物,尤其是,二硫键183-190对于WntLys182和FZGlu64之间的远程盐桥相互作用建立非常重要,导致棕榈酰基基团适当定位到FZ,提示这种二硫键在Wnt-FZ复合物形成中的重要作用。一起,我们的发现为拇指定位的二硫化物如何有助于Wnt-FZ复合物的形成提供了新的见解,结构动力学,和稳定性,在针对Wnt信号传导的药物设计和开发中引入二硫化物183-190作为目标的重要元件。
Wnts are lipid-modified proteins rich in cysteine, regulating developmental processes, and are involved in various pathological conditions. Wnts structure resembles a hand, with a palmitoleylated thumb and an index finger-like domain interacting with
frizzled (FZ) receptors. Previous research shows the palmitoleyl group and the disulfides importance in Wnt folding, secretion, and function, but the structural basis is not fully understood. Here, we utilized classical molecular dynamics simulation (800-ns in total) to investigate how the thumb palmitoleyl and its close conserved disulfides (183-190, 181-195) regulated Wnt-FZ interaction and structural dynamics. Using Steered molecular dynamics experiment followed by a relaxing procedure, we also explored if these disulfides are important in Wnt-FZ complex formation. According to our results, the palmitoleyl group contributes significantly to stabilize Wnt-FZ interaction, and the disulfides modulate this contribution. We also demonstrated that disulfide 183-190 regulates the Wnt thumb fluctuation, hydrogen bond network, and secondary structure. The DCCM analysis depicted disulfide 183-190 roles in regulating native-like collective movement in the palmitoleylated loop, which changed after this disulfide removal. The pulling-relaxing experiment showed that both the disulfides, and especially, the disulfide 183-190, are highly important for long-range salt-bridge interaction establishment between Wnt Lys182 and FZ Glu64, led palmitoleyl group appropriate positioning to FZ, suggested this disulfide essential role in Wnt-FZ complex formation. Together, our findings provide new insights to how thumb-positioned disulfides contribute to Wnt-FZ complex formation, structural dynamics, and stability, introducing disulfide 183-190 as a consequential element to target in drug design and development against Wnt signalling.