Aggregation-resistant domain antibodies selected on phage by heat denaturation
Aggregation-resistant domain antibodies selected on phage by heat denaturation
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DOI：
10.1038/nbt1000
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发表时间：
2004-09-01
影响因子：
46.9
通讯作者：
Winter, G
中科院分区：
文献类型：
--
作者：
Jespers, L;Schon, O;Winter, G
我们描述了一种通过热变性来选择抗聚集蛋白的方法。抗体重链可变结构域（dAb）就说明了这一点，它们容易聚集（1，2）。dAb在丝状噬菌体的感染尖端多价展示，并瞬时加热以诱导解折叠并促进dAb的聚集。冷却后，选择dAb与蛋白A（这些折叠dAb共有的配体）结合。由此相对于不可逆地解折叠的噬菌体展示dAb富集。从噬菌体dAb的库中，选择后表征了六种dAb;它们都抵抗聚集，并且是可溶的，在细菌中良好表达，并且可以以良好的产量纯化。该方法应可用于制备抗聚集蛋白质，并有助于鉴定促进或阻止蛋白质聚集的特征，包括那些导致错误折叠疾病的特征（3，4）。
We describe a method for selecting aggregation-resistant proteins by heat denaturation. This is illustrated with antibody heavy chain variable domains (dAbs), which are prone to aggregate(1,2). The dAbs were displayed multivalently at the infective tip of filamentous bacteriophage, and heated transiently to induce unfolding and to promote aggregation of the dAbs. After cooling, the dAbs were selected for binding to protein A (a ligand common to these folded dAbs). Phage displaying dAbs that unfold reversibly were thereby enriched with respect to those that do not. From a repertoire of phage dAbs, six dAbs were characterized after selection; they all resisted aggregation, and were soluble, well expressed in bacteria and could be purified in good yields. The method should be useful for making aggregation-resistant proteins and for helping to identify features that promote or prevent protein aggregation, including those responsible for misfolding diseases(3,4).