Molecular Evolution of Antiparathion Nanobody with Enhanced Sensitivity and Specificity Based on Structural Analysis
Molecular Evolution of Antiparathion Nanobody with Enhanced Sensitivity and Specificity Based on Structural Analysis
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DOI：
10.1021/acs.jafc.3c05176
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发表时间：
2023-09-28
影响因子：
6.1
通讯作者：
Wang，Hong
中科院分区：
文献类型：
--
作者：
Li，Jia-Dong;Shen，Xing;Wang，Hong
Nanobody (Nb) has gained significant attention in immunoassays owing to its numerous advantages, particularly its ease of molecular evolution. However, the limited understanding of how high sensitivity and specificity attained for antihapten Nbs hamper the development of high-performance Nbs. Herein, the antiparathion Nb (Nb9) we prepared previously was chosen as the model, and an approach based on X-ray crystallography, molecular docking, and rational site-directed saturation mutation for constructing a rapid and effective platform for nanobody evolution was described. Based on the structural analysis, two mutants, namely Nb-D5 (IC50= 2.4 ± 0.2 ng/mL) and Nb-D12 (IC50= 2.7 ± 0.1 ng/mL), were selected out from a six-sites directed saturation mutation library, 3.5-fold and 3.1-fold sensitivity enhancement over Nb9 to parathion, respectively. Besides, Nb-D12 exhibited improved sensitivity for quinalphos, triazophos, and coumaphos (5.4–35.4 ng/mL), indicating its broader detection potential. Overall, our study advances an effective strategy for the future rational evolution of Nbs with desirable performance.
