人类癌毒素M受体亚基，通常称为癌毒素M受体（OSMR），是一种细胞表面蛋白质，属于I型细胞因子受体家族。它在多种癌症中高度表达，是一个潜在的治疗靶点。从结构上讲，OSMR由三个主要域组成：细胞外、跨膜和胞质域。细胞外结构域进一步包括四个III型纤维连接蛋白亚域。这些III型纤维连接蛋白域的功能相关性尚不清楚，我们非常有兴趣了解它们在OSMR介导的与其他致病蛋白的相互作用中的作用。hOSMR的四个III型纤维连接蛋白域是用pUNO1-hOSMR构建物作为模板进行PCR扩增的。扩增产物的分子大小通过琼脂糖凝胶电泳进行确认。然后将片段克隆到一个含有N末端标签为GST的pGEX4T3载体中。通过限制性酶切鉴定含有域插入的阳性克隆并在大肠杆菌Rosetta（DE3）细胞中过表达。过表达的最佳条件为1 mM IPTG和37℃的孵育温度。纤维连接蛋白域的过表达通过SDS-PAGE确认，并使用谷胱甘肽琼脂糖糖珠进行三次重复步骤的亲和纯化。通过SDS-PAGE和Western blot分析分离的纯度表明它们恰好在与其对应的分子量的单一明显波段处。在这项研究中，我们成功克隆、表达和纯化了hOSMR的四个III型纤维连接蛋白亚域。©2023.作者（们），在Springer Nature B.V.的独家许可下。

The human oncostatin M receptor subunit , commonly known as the oncostatin M receptor (OSMR), is a cell surface protein and belongs to the family of type I cytokine receptors. It is highly expressed in several cancers and is a potential therapeutic target. Structurally, OSMR consists of three major domains: the extracellular, transmembrane, and cytoplasmic domains. The extracellular domain further comprises four Type III fibronectin subdomains. The functional relevance of these type III fibronectin domains is not known yet, and it is of great interest to us to understand their role in OSMR-mediated interactions with other oncogenic proteins.The four type III fibronectin domains of hOSMR were amplified by PCR using the pUNO1-hOSMR construct as a template. The molecular size of the amplified products was confirmed by agarose gel electrophoresis. The amplicons were then cloned into a pGEX4T3 vector containing GST as an N-terminal tag. Positive clones with domain inserts were identified by restriction digestion and overexpressed in E. coli Rosetta (DE3) cells. The optimum conditions for overexpression were found to be 1 mM IPTG and an incubation temperature of 37 °C. The overexpression of the fibronectin domains was confirmed by SDS-PAGE, and they are affinity purified by using glutathione agarose beads in three repetitive steps. The purity of the isolated domains analyzed by SDS-PAGE and western blotting showed that they were exactly at their corresponding molecular weights as a single distinct band.In this study, we have successfully cloned, expressed, and purified four Type III fibronectin subdomains of hOSMR.© 2023. The Author(s), under exclusive licence to Springer Nature B.V.