Forage Biotechnology, AgResearch Grasslands, Palmerston North, New Zealand.
J Biotechnol. 2012 Nov 15;161(4):407-13. doi: 10.1016/j.jbiotec.2012.07.195. Epub 2012 Aug 16.
A variety of single-chain variable fragments (scFv) that had been previously developed to the surface epitopes of infective Trichostrongylus colubriformis L3 pathogenic gut nematodes of sheep were fused to a trimeric version of polyoleosin (three head-to-tail repeats of oleosin) and expressed in planta under the control of an Arabidopsis oleosin promoter. The fusion products were found to accumulate in oil bodies (OBs) at the range of 0.25-0.9% of the total seed protein which is comparable with the main 18 kDa isoform of Arabidopsis seed oleosin. Immunofluorescence microscopy and immuno-binding were used to demonstrate that it is possible to both purify the recombinant protein via enrichment for OBs as well as use the OBs emulsion to deliver functional recombinant scFv. This work presents a novel fusion strategy platform to boost the productivity and simplify the delivery of recombinant single chain antibodies and other like proteins.
已开发出多种针对绵羊感染性旋毛线虫 L3 致病肠道线虫表面表位的单链可变片段 (scFv)，将其与三聚体聚油蛋白 (油蛋白的三个头尾重复) 融合，并在拟南芥油蛋白启动子的控制下在植物中表达。融合产物被发现以 0.25-0.9%的总种子蛋白的形式积累在油体 (OBs) 中，这与拟南芥种子油蛋白的主要 18 kDa 同工型相当。免疫荧光显微镜和免疫结合用于证明可以通过富集 OBs 来纯化重组蛋白，并且可以使用 OBs 乳液来输送功能性重组 scFv。这项工作提出了一种新的融合策略平台，以提高重组单链抗体和其他类似蛋白的产量并简化其传递。
