Keywords

Linker Polypeptide, Ietd, Eeed, F2A, Caspase Activity

Abstract

IL-24 and smac are important candidate genes in the study of multi-gene therapy targeting cancer. In the process of constructing the il-24 and smac dual gene expression vector, it is very critical to select a stable and efficient linker. Three different linker polypeptide sequences ietd, eeed, and f2a were used to construct three eukaryotic expression plasmids pcdna3.1(+)-il-24-ietd-smac, pcdna3.1()-il-24-eeed-smac, pcdna3.1()-il-24-f2a-smac, and combined with 5-fluorouracil (5-fu) treatment to study the fusion protein cleavage effect mediated by each linker. The above experimental results show that among the three il-24-linker-smac dual-gene expression vectors, the f2a linker has the best cutting effect and is positively correlated with caspase activity. The fusion proteins mediated by the ietd and eeed linkers have a certain degree of shearing, but the ietd/eeed polypeptide affects the half-life of the upstream il-24 protein and accelerates the ubiquitination and degradation of the il-24 protein after shearing. This study provides a design reference for constructing dual-gene or multi-gene vectors for cancer-targeting applications.

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Original research done by Li Hongyan, Zhou Zhiming, Pan Qiang, Yang Dongmei, Gu Jinfa, Shen Jiani, Wang Shibing, Zhang Kangjian

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