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首页> 外文期刊>Journal of Molecular Biology >AFFINITY SELECTIVE ISOLATION OF LIGANDS FROM PEPTIDE LIBRARIES THROUGH DISPLAY ON A LAC REPRESSOR HEADPIECE DIMER
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AFFINITY SELECTIVE ISOLATION OF LIGANDS FROM PEPTIDE LIBRARIES THROUGH DISPLAY ON A LAC REPRESSOR HEADPIECE DIMER

机译：通过在Lac阻遏物头二聚体上显示的肽库中的脂质进行亲和性选择性分离

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摘要

DNA binding by the Escherichia coli lac repressor is mediated by the similar to 60 amino acid residue 'headpiece' domain. The dimer of headpiece domains that binds to the lac operator is normally formed by association of the much larger similar to 300 amino acid residue C-terminal domain. We have used in vitro selection to isolate 'headpiece dimer' molecules containing two headpiece domains connected via a short peptide linker. These proteins bind plasmid molecules with sufficient stability to allow association of a peptide epitope displayed at the C terminus of the headpiece dimer with the plasmid encoding that peptide. Libraries of peptides displayed on the C terminus of a headpiece dimer can be screened for specific receptor ligands by affinity enrichment of peptide-headpiece dimer-plasmid complexes using an immobilized receptor. After each round of enrichment, transformation of E. coli with recovered plasmids permits amplification of the selected population. After several rounds of enrichment, sequencing of individual clones reveals the structure of the selected peptides. Headpiece dimer libraries allow selection of peptide ligands of higher average affinity than similar libraries based on the intact lac repressor. Interestingly, the presence of the lac operator is not required for plasmid binding by the headpiece dimer protein. (C) 1996 Academic Press Limited [References: 59]
机译：大肠杆菌lac阻遏物与DNA的结合是由类似于60个氨基酸残基的“头部”结构域介导的。与lac操纵子结合的头盔结构域的二聚体通常是通过与300个氨基酸残基C端结构域相似的更大的缔合而形成的。我们已经使用体外选择来分离包含两个通过短肽接头连接的头域的“头二聚体”分子。这些蛋白质以足够的稳定性结合质粒分子，以使在头饰二聚体的C末端显示的肽表位与编码该肽的质粒缔合。可以使用固定的受体通过亲和富集肽-头戴二聚体-质粒复合物，筛选显示在头戴二聚体C末端的肽文库中的特定受体配体。在每一轮富集之后，用回收的质粒转化大肠杆菌可以扩增选定的群体。经过数轮富集后，对单个克隆进行测序可揭示所选肽的结构。头戴二聚体文库允许选择比基于完整lac阻遏物的类似文库更高的平均亲和力的肽配体。有趣的是，通过头件二聚体蛋白结合质粒不需要lac操纵子的存在。 （C）1996 Academic Press Limited [参考号：59]

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