Virology Laboratory, Universidad Javeriana, Bogotá DC, Colombia.
Nutrition and Biochemistry Department, Science Faculty, Universidad Javeriana, Bogotá DC, Colombia.
PLoS One. 2018 Jun 21;13(6):e0199397. doi: 10.1371/journal.pone.0199397. eCollection 2018.
The envelope glycoprotein 51 (gp51) is essential for bovine leukaemia virus (BLV) entry to bovine B-lymphocytes. Although the bovine adaptor protein 3 complex subunit delta-1 (boAP3D1) has been proposed as the potential receptor, the specific ligand-receptor interaction has not yet been completely defined and boAP3D1 receptor and gp51 3D structures have not been determined. This study was thus aimed at a functional annotation of boAP3D1 cellular adaptor protein and BLV gp51 and, proposing a reliable model for gp51-AP3D1 interaction using bioinformatics tools. The boAP3D1 receptor interaction patterns were calculated based on models of boAP3D1 receptor and gp51 complexes' 3D structures, which were constructed using homology techniques and data-driven docking strategy. The results showed that the participation of 6 key amino acids (aa) on gp51 (Asn170, Trp127, His115, Ala97, Ser98 and Glu128) and 4 aa on AP3D1 (Lys925, Asp807, Asp695 and Arg800) was highly probable in the interaction between gp51 and BLVR domains. Three gp51 recombinant peptides were expressed and purified to validate these results: the complete domain (rgp51), the N-terminal portion (rNgp51) and the C-terminal fragment (rCgp51); and binding assays to Madin-Darby bovine kidney (MDBK) cells were then carried out with each recombinant. It was found that rNgp51 preferentially bound to MDBK cells, suggesting this domain's functional role during invasion. The rNgp51-MDBK cell interaction was sensitive to trypsin (98% reduction) and chymotrypsin treatment (80% reduction). These results highlighted that the N-terminal portion of gp51 interacted in vitro with the AP3D1 receptor and provides a plausible in silico interaction model.
包膜糖蛋白 51（gp51）对于牛白血病病毒（BLV）进入牛 B 淋巴细胞是必不可少的。虽然已经提出牛衔接蛋白 3 复合物亚基 δ-1（boAP3D1）是潜在的受体，但特定的配体-受体相互作用尚未完全定义，并且 boAP3D1 受体和 gp51 3D 结构尚未确定。因此，本研究旨在对 boAP3D1 细胞衔接蛋白和 BLV gp51 进行功能注释，并使用生物信息学工具提出 gp51-AP3D1 相互作用的可靠模型。基于同源技术和数据驱动对接策略构建的 boAP3D1 受体和 gp51 复合物 3D 结构模型，计算了 boAP3D1 受体的相互作用模式。结果表明，gp51 上的 6 个关键氨基酸（aa）（Asn170、Trp127、His115、Ala97、Ser98 和 Glu128）和 AP3D1 上的 4 个 aa（Lys925、Asp807、Asp695 和 Arg800）在 gp51 和 BLVR 结构域之间的相互作用中高度可能。表达和纯化了 3 个 gp51 重组肽以验证这些结果：完整结构域（rgp51）、N 端部分（rNgp51）和 C 端片段（rCgp51）；并随后对每个重组蛋白进行了与 Madin-Darby 牛肾（MDBK）细胞的结合测定。结果发现 rNgp51 优先与 MDBK 细胞结合，这表明该结构域在入侵过程中具有功能作用。rNgp51-MDBK 细胞相互作用对胰蛋白酶（98%降低）和糜蛋白酶处理（80%降低）敏感。这些结果突出表明 gp51 的 N 端部分在体外与 AP3D1 受体相互作用，并提供了一个合理的计算机模拟相互作用模型。
