Hao Junfang, Wang Fangyu, Xing Guangxu, Liu Yunchao, Deng Ruiguang, Zhang Hao, Cheng Anchun, Zhang Gaiping
Research Center of Avian Diseases, College of Veterinary Medicine, Sichuan Agricultural University, Chengdu, China.
Henan Key Laboratory for Animal Immunology, Henan Academy of Agricultural Sciences, Zhengzhou, China.
PeerJ. 2019 Dec 5;7:e8132. doi: 10.7717/peerj.8132. eCollection 2019.
Affinity peptides, as a core part of affinity chromatography, play an important role in the purification of target molecules.
Here we describe the use of molecular docking technology for virtual screening of affinity peptides that specifically recognize the PCV2 Cap protein for the first time. Thirteen candidate peptides with high scores were obtained and then further characterized. Experimentally, the affinity and sensitivity of the peptides studied were identified by ELISA and LSPR, respectively. In order to investigate the purification effect of a selected peptide (L11) for the recombinant PCV2 Cap protein, it was coupled to NHS agarose magnetic beads as an affinity adsorbent (NaMB-L11); and the ligand density of the affinity adsorbent and pH value in the purification of the recombinant PCV2 Cap protein were optimized.
Our data showed that the peptide L11- DYWWQSWE has the smallest K = 103 nM with higher specificity for PCV2 Cap protein recognition. The NaMB-L11 affinity adsorbent yielded a purified Cap sample with 98% purity at 90% recovery in a single step.
Based on the structure, we obtained a high affinity peptide L11 binding to the PCV2 Cap protein by molecular docking technology. It not only provides a theoretical basis for the design of PCV2 Cap affinity peptide, but a new method for the purification of the PCV2 Cap protein.
亲和肽作为亲和色谱的核心部分，在目标分子的纯化中发挥着重要作用。
在此，我们首次描述了使用分子对接技术对特异性识别猪圆环病毒2型（PCV2）Cap蛋白的亲和肽进行虚拟筛选。获得了13个高分候选肽，然后对其进行进一步表征。在实验中，分别通过酶联免疫吸附测定（ELISA）和局域表面等离子体共振（LSPR）确定所研究肽的亲和力和灵敏度。为了研究所选肽（L11）对重组PCV2 Cap蛋白的纯化效果，将其偶联到N - 羟基琥珀酰亚胺（NHS）琼脂糖磁珠上作为亲和吸附剂（NaMB - L11）；并对亲和吸附剂的配体密度和重组PCV2 Cap蛋白纯化过程中的pH值进行了优化。
我们的数据表明，肽L11 - DYWWQSWE的最小解离常数K = 103 nM，对PCV2 Cap蛋白识别具有更高的特异性。NaMB - L11亲和吸附剂在一步操作中可获得纯度为98%、回收率为90%的纯化Cap样品。
基于结构，我们通过分子对接技术获得了一种与PCV2 Cap蛋白结合的高亲和力肽L11。它不仅为PCV2 Cap亲和肽的设计提供了理论依据，也为PCV2 Cap蛋白的纯化提供了一种新方法。
