Departamento de Medicina Molecular y Bioprocesos, Instituto de Biotecnología, Universidad Nacional Autónoma de México, Apartado Postal 510-3, Cuernavaca, Morelos 62210, USA.
J Biol Chem. 2011 Jun 10;286(23):20892-900. doi: 10.1074/jbc.M111.238410. Epub 2011 Apr 13.
It has previously been reported that several single-chain antibody fragments of human origin (scFv) neutralize the effects of two different scorpion venoms through interactions with the primary toxins of Centruroides noxius Hoffmann (Cn2) and Centruroides suffusus suffusus (Css2). Here we present the crystal structure of the complex formed between one scFv (9004G) and the Cn2 toxin, determined in two crystal forms at 2.5 and 1.9 Å resolution. A 15-residue span of the toxin is recognized by the antibody through a cleft formed by residues from five of the complementarity-determining regions of the scFv. Analysis of the interface of the complex reveals three features. First, the epitope of toxin Cn2 overlaps with essential residues for the binding of β-toxins to its Na(+) channel receptor site. Second, the putative recognition of Css2 involves mainly residues that are present in both Cn2 and Css2 toxins. Finally, the effect on the increase of affinity of previously reported key residues during the maturation process of different scFvs can be inferred from the structure. Taken together, these results provide the structural basis that explain the mechanism of the 9004G neutralizing activity and give insight into the process of directed evolution that gave rise to this family of neutralizing scFvs.
先前有报道称，几种源自人类的单链抗体片段（scFv）通过与 Centruroides noxius Hoffmann（Cn2）和 Centruroides suffusus suffusus（Css2）的主要毒素相互作用，中和了两种不同蝎毒的作用。在这里，我们展示了与 Cn2 毒素形成复合物的一个 scFv（9004G）的晶体结构，该结构以 2.5 和 1.9 Å 的分辨率在两种晶体形式下确定。通过 scFv 的五个互补决定区的残基形成的裂缝，抗体识别毒素的 15 个残基跨度。对复合物界面的分析揭示了三个特征。首先，Cn2 毒素的表位与 β-毒素与其 Na(+)通道受体结合的必需残基重叠。其次，Css2 的假定识别主要涉及存在于 Cn2 和 Css2 毒素中的残基。最后，结构可以推断出在不同 scFv 的成熟过程中关键残基亲和力增加的先前报道的效果。综上所述，这些结果提供了解释 9004G 中和活性的机制的结构基础，并深入了解了产生这种中和 scFv 家族的定向进化过程。
