Rapid and Direct VHH and Target Identification by Staphylococcal Surface Display Libraries.
Rapid and Direct VHH and Target Identification by Staphylococcal Surface Display Libraries.
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DOI：
10.3390/ijms18071507
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发表时间：
2017-07-12
影响因子：
5.6
通讯作者：
Cavallari M
中科院分区：
文献类型：
--
作者：
Cavallari M
在没有至少一种相互作用配偶体的先验知识的情况下，特异性抗体及其靶抗原的无偏倚和同时鉴定是困难的。用抗原的复杂混合物如整个生物体和组织提取物（包括肿瘤提取物）进行免疫引起高度多样化的免疫应答。在这样的应答期间，产生针对混合物中的多种表位的抗体。在这里，我们提出了一个表面展示设计，适合于同时识别骆驼单域抗体和它们的目标。将来自骆驼科动物仅重链抗体（VHH）的单结构域抗原识别片段的免疫文库使用其内源性管家分选酶连接至革兰氏阳性金黄色葡萄球菌的肽聚糖。分选酶转肽反应将VHH共价连接至细菌肽聚糖。反应的可逆性质允许从细菌表面回收VHH并在下游应用中使用VHH。这些葡萄球菌表面展示文库用于通过免疫沉淀（IP）快速鉴定VHH及其靶标。我们的新型细菌表面展示平台在苛刻的筛选条件下是稳定的，允许快速的靶标鉴定，并且容易允许回收展示的VHH用于下游分析。
Unbiased and simultaneous identification of a specific antibody and its target antigen has been difficult without prior knowledge of at least one interaction partner. Immunization with complex mixtures of antigens such as whole organisms and tissue extracts including tumoral ones evokes a highly diverse immune response. During such a response, antibodies are generated against a variety of epitopes in the mixture. Here, we propose a surface display design that is suited to simultaneously identify camelid single domain antibodies and their targets. Immune libraries of single-domain antigen recognition fragments from camelid heavy chain-only antibodies (VHH) were attached to the peptidoglycan of Gram-positive Staphylococcus aureus employing its endogenous housekeeping sortase enzyme. The sortase transpeptidation reaction covalently attached the VHH to the bacterial peptidoglycan. The reversible nature of the reaction allowed the recovery of the VHH from the bacterial surface and the use of the VHH in downstream applications. These staphylococcal surface display libraries were used to rapidly identify VHH as well as their targets by immunoprecipitation (IP). Our novel bacterial surface display platform was stable under harsh screening conditions, allowed fast target identification, and readily permitted the recovery of the displayed VHH for downstream analysis.
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影响因子：
14.8
作者：
通讯作者：
--
影响因子：
6.4
作者：
Hay ID;Du J;Reyes PR;Rehm BH
通讯作者：
Rehm BH
影响因子：
3.6
作者：
Harmsen, MM;Ruuls, RC;de Geus, B
通讯作者：
de Geus, B
影响因子：
4.8
作者：
Govaert, Jochen;Pellis, Mireille;Saerens, Dirk
通讯作者：
Saerens, Dirk
DOI：
10.1111/j.1432-1033.1984.tb07931.x
发表时间：
1984-01-01
期刊：
EUROPEAN JOURNAL OF BIOCHEMISTRY
影响因子：
--
作者：
GUSS, B;UHLEN, M;SJODAHL, J
通讯作者：
SJODAHL, J