Departamento de Bioquímica e Biologia Molecular, Universidade Federal do Ceará, Campus do Pici, 60440970 Fortaleza, Ceará, Brazil.
Universidade Federal do Rio Grande do Norte, Escola Agrícola de Jundiaí, Distrito de Jundiaí, 59280000 Macaíba, Rio Grande do Norte, Brazil.
Int J Biol Macromol. 2020 Aug 1;156:1-9. doi: 10.1016/j.ijbiomac.2020.04.009. Epub 2020 Apr 7.
Lectins from Diocleinae subtribe species (family Leguminosae) are of special interest since they present a wide spectrum of biological activities, despite their high structural similarity. During their synthesis in plant cells, these proteins undergo post-translational processing resulting in the formation of three chains (α, β, γ), which constitute the lectins' subunits. Furthermore, such wild-type proteins are presented as isolectins or with different combinations of these chains, which undermine their biotechnological potential. Thus, the present study aimed to produce a recombinant form of the lectin from Dioclea sclerocarpa seeds (DSL), exclusively constituted by α-chain. The recombinant DSL (rDSL) was successfully expressed in E. coli BL21 (DE3) and purified by affinity chromatography (Sephadex G-50), showing a final yield of 74 mg of protein per liter of culture medium and specificity for D-mannose, α-methyl-mannoside and melibiose, unlike the wild-type protein. rDSL presented an effective vasorelaxant effect in rat aortas up to 100% and also interacted with glioma cells C6 and U87. Our results demonstrated an efficient recombinant production of rDSL in a bacterial system that retained some biochemical properties of the wild-type protein, showing wider versatility in sugar specificities and better efficacy in its activity in the biological models evaluated in this work.
Diocleinae 亚族（豆科）物种的凝集素特别有趣，因为它们具有广泛的生物活性，尽管它们的结构高度相似。在植物细胞中合成时，这些蛋白质会经历翻译后加工，形成由三个链（α、β、γ）构成的亚基，从而构成凝集素。此外，这些野生型蛋白质通常表现为同工凝集素或具有这些链的不同组合，这限制了它们的生物技术潜力。因此，本研究旨在从 Dioclea sclerocarpa 种子中产生一种重组形式的凝集素（DSL），仅由 α 链组成。重组 DSL（rDSL）在大肠杆菌 BL21（DE3）中成功表达，并通过亲和层析（Sephadex G-50）进行纯化，每升培养基的最终产量为 74 毫克蛋白质，并且对 D-甘露糖、α-甲基甘露糖苷和棉子糖具有特异性，这与野生型蛋白质不同。rDSL 在大鼠主动脉中表现出高达 100%的有效血管舒张作用，并且还与神经胶质瘤细胞 C6 和 U87 相互作用。我们的结果表明，rDSL 在细菌系统中的重组生产是有效的，该系统保留了野生型蛋白的一些生化特性，在评估的生物模型中表现出更广泛的糖特异性和更好的活性。
