Departamento de Fisiologia e Biofísica, Instituto de Ciências Biológicas, Universidade Federal de Minas Gerais, Av Antônio Carlos 6627, Pampulha, 31270-901 Belo Horizonte, Minas Gerais, Brazil.
Metallomics. 2012 May;4(5):433-40. doi: 10.1039/c2mt00176d. Epub 2012 Mar 27.
It has been reported recently that Sb(III) competes with Zn(II) for its binding to the CCHC zinc finger domain of the HIV-1 NCp7 protein, suggesting that zinc finger proteins may be molecular targets for antimony-based drugs. Here, the interaction of Sb(III) with a CCCH zinc finger domain, which is considered to play a crucial role in the biology of kinetoplastid protozoa, has been characterized for the first time. The binding characteristics of Sb(III) were compared between a CCCH-type peptide derived from a kinetoplastid protein and two different CCHC-type zinc finger peptides. The formation of 1 : 1 Zn-peptide and Sb-peptide complexes from the different peptides was demonstrated using circular dichroism, UV absorption, fluorescence spectroscopies and ESI-MS. Titration of the Zn-peptide complexes with SbCl(3) was performed at pH 6 and 7, exploiting the intrinsic fluorescence of the peptides. The differential spectral characteristics of the peptides allowed for competition experiments between the different peptides for binding of Zn(II). The present study establishes that Sb(III) more effectively displaces Zn(II) from the CCCH peptide than CCHC ones, as a result of both the greater stability of the Sb-CCCH complex (compared to Sb-CCHC complexes) and the lower stability of the Zn-CCCH complex (compared to Zn-CCHC complexes). Comparison of the binding characteristics of Sb(III) or Zn(II) between the CCHC-type peptides with different amino acid sequences supports the model that not only the conserved zinc finger motif, but also the sequence of non-conserved amino acids determines the binding affinity of Sb(III) and Zn(II). These data suggest that the interaction of Sb(III) with CCCH-type zinc finger proteins may modulate, or even mediate, the pharmacological action of antimonial drugs.
最近有报道称，Sb(III)与 Zn(II)竞争与 HIV-1 NCp7 蛋白的 CCHC 锌指结构域结合，这表明锌指蛋白可能是基于锑的药物的分子靶标。在这里，首次对 CCCH 锌指结构域与 Sb(III)的相互作用进行了表征，该结构域被认为在动基体原生动物的生物学中发挥着关键作用。比较了来自动基体蛋白的 CCCH 型肽和两种不同的 CCHC 型锌指肽之间 Sb(III)的结合特性。利用圆二色性、紫外吸收、荧光光谱和 ESI-MS 证明了不同肽段形成 1:1 Zn-肽和 Sb-肽复合物。在 pH 6 和 7 下，用 SbCl3 滴定 Zn-肽配合物，利用肽的固有荧光。不同肽的差异光谱特性允许进行不同肽之间竞争结合 Zn(II)的实验。本研究确立了 Sb(III)比 CCHC 更有效地从 CCCH 肽中置换 Zn(II)，这是由于 Sb-CCCH 配合物的稳定性（与 Sb-CCHC 配合物相比）更高，以及 Zn-CCCH 配合物的稳定性（与 Zn-CCHC 配合物相比）更低。比较不同氨基酸序列的 CCHC 型肽中 Sb(III)或 Zn(II)的结合特性支持这样的模型，即不仅保守的锌指基序，而且非保守氨基酸序列决定 Sb(III)和 Zn(II)的结合亲和力。这些数据表明，Sb(III)与 CCCH 型锌指蛋白的相互作用可能调节甚至介导锑药物的药理作用。
