Plants are economical and sustainable factories for the production of recombinant proteins. Currently, numerous proteins produced using different plant-based systems with applications as cosmetic and tissue culture ingredients, research and diagnostic reagents, and industrial enzymes are marketed worldwide. In this study, we aimed to demonstrate the usefulness of a plant-based system to synthesize a single-chain antibody (scFv)—elastin-like polypeptide (ELP) fusion to be applied as an affinity precipitation reagent of the difficult to produce recombinant proteins. We used the human tissue transglutaminase (TG2), the main celiac disease autoantigen, as a proof of concept. We cloned a TG2-specific scFv and fused it to a short hydrophobic ELP tag. The anti-TG2-scFv-ELP was produced in Nicotiana benthamiana and was efficiently recovered by an inverse transition cycling procedure improved by coaggregation with bacteria-made free ELP. Finally, the scFv-ELP was used to purify both plant-synthesized human TG2 and also Caco-2-TG2. In conclusion, this study showed for the first time the usefulness of a plant-based expression system to produce an antibody-ELP fusion designed for the purification of low-yield proteins.

中文翻译：

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本氏烟草类弹性蛋白多肽融合单链抗体片段的合成及其在难生产蛋白亲和沉淀中的应用


植物是生产重组蛋白的经济且可持续的工厂。目前，使用不同植物系统生产的多种蛋白质在全球范围内销售，这些蛋白质可用作化妆品和组织培养成分、研究和诊断试剂以及工业酶。在这项研究中，我们旨在证明基于植物的系统合成单链抗体（scFv）-弹性蛋白样多肽（ELP）融合物的有用性，该融合物可用作难以生产的重组蛋白的亲和沉淀试剂。我们使用人体组织转谷氨酰胺酶 (TG2)（主要的乳糜泻自身抗原）作为概念证明。我们克隆了 TG2 特异性 scFv 并将其融合到短疏水性 ELP 标签上。抗 TG2-scFv-ELP 在本塞姆氏烟草中产生，并通过与细菌产生的游离 ELP 共聚改进的逆转变循环程序有效回收。最后，使用 scFv-ELP 纯化植物合成的人 TG2 和 Caco-2-TG2。总之，这项研究首次表明基于植物的表达系统可用于生产专为纯化低产蛋白而设计的抗体-ELP 融合体。