Fv-clasp: An Artificially Designed Small Antibody Fragment with Improved Production Compatibility, Stability, and Crystallizability
Fv-clasp: An Artificially Designed Small Antibody Fragment with Improved Production Compatibility, Stability, and Crystallizability
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DOI：
10.1016/j.str.2017.08.011
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发表时间：
2017-10-03
期刊：
影响因子：
5.7
通讯作者：
Takagi, Junichi
中科院分区：
文献类型：
--
作者：
Arimori, Takao;Kitago, Yu;Takagi, Junichi
抗体片段经常被用作“结晶伴侣”，以帮助复杂的大分子的结构分析，否则这些大分子是抗结晶的，但是常规的片段形式还没有被设计用于这种特定的应用。通过将来自人Mst 1激酶的SARAH结构域的反平行卷曲螺旋结构融合到抗体的可变区，我们成功地产生了类似于37 kDa的新型嵌合抗体片段，称为“Fv-扣”，其表现出优异的结晶相容性，同时保持了原始IgG分子的结合能力。“扣”和Fv底部的工程化二硫键抑制了片段的内部移动性并屏蔽了疏水残基，这可能有助于Fv-扣的高热稳定性和可结晶性。最后，Fv-钩状抗体显示出优于常规Fab片段的上级“陪伴”活性，并促进了整联蛋白α 6 β 1的胞外域片段的结构测定。
Antibody fragments are frequently used as a "crystallization chaperone'' to aid structural analysis of complex macromolecules that are otherwise crystallization resistant, but conventional fragment formats have not been designed for this particular application. By fusing an anti-parallel coiled-coil structure derived from the SARAH domain of human Mst1 kinase to the variable region of an antibody, we succeeded in creating a novel chimeric antibody fragment of similar to 37 kDa, termed "Fv-clasp,'' which exhibits excellent crystallization compatibility while maintaining the binding ability of the original IgG molecule. The "clasp'' and the engineered disulfide bond at the bottom of the Fv suppressed the internal mobility of the fragment and shielded hydrophobic residues, likely contributing to the high heat stability and the crystallizability of the Fv-clasp. Finally, Fv-clasp antibodies showed superior "chaperoning'' activity over conventional Fab fragments, and facilitated the structure determination of an ectodomain fragment of integrin alpha 6 beta 1.