The Mechanochemistry of a Structural Zinc Finger.
The Mechanochemistry of a Structural Zinc Finger.
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DOI：
10.1021/acs.jpclett.5b01371
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发表时间：
2015-08
期刊：
影响因子：
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通讯作者：
Judit Perales-Calvo;Ainhoa Lezamiz;S. Garcia-Manyes
中科院分区：
文献类型：
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作者：
Judit Perales-Calvo;Ainhoa Lezamiz;S. Garcia-Manyes
锌指是非常普遍的结构基序，可为蛋白质提供稳定性，从而有助于其正确折叠。尽管 ZnCys4 中心具有很高的热力学稳定性，但它们的动力学性质表现出显着的不稳定性。在这里，我们将蛋白质工程与单分子力谱原子力显微镜 (AFM) 相结合，揭示了单个 Zn-S 键令人惊讶的机械不稳定性 (∼90 pN)，这些键形成嵌入多域 DnaJ 分子伴侣结构中的两个等效锌指基序。锌配位残基内的合理突变能够直接识别调节锌结合（需要所有四个半胱氨酸存在）和二硫键形成之间相互作用的化学决定因素。最后，我们的观察表明，与疏水性短肽的结合大大增加了 DnaJ 的机械稳定性。总而言之，我们的实验方法为控制单个自然产生的锌指的纳米力学的精细化学机制提供了详细的原子论前景。
Zinc fingers are highly ubiquitous structural motifs that provide stability to proteins, thus contributing to their correct folding. Despite the high thermodynamic stability of the ZnCys4 centers, their kinetic properties display remarkable lability. Here, we use a combination of protein engineering with single molecule force spectroscopy atomic force microscopy (AFM) to uncover the surprising mechanical lability (∼90 pN) of the individual Zn-S bonds that form the two equivalent zinc finger motifs embedded in the structure of the multidomain DnaJ chaperone. Rational mutations within the zinc coordinating residues enable direct identification of the chemical determinants that regulate the interplay between zinc binding-requiring the presence of all four cysteines-and disulfide bond formation. Finally, our observations show that binding to hydrophobic short peptides drastically increases the mechanical stability of DnaJ. Altogether, our experimental approach offers a detailed, atomistic vista on the fine chemical mechanisms that govern the nanomechanics of individual, naturally occurring zinc finger.