Cell-free protein synthesis of CD1E and B2M protein and in vitro interaction

文献类型： 外文期刊

第一作者： Tang, Yajie

作者： Tang, Yajie;Ma, Shengming;Wu, Yinrong;Chen, Siyang;Wang, Yao;Jiang, Lele;Lin, Sen;Ma, Lisong;Wang, Zaihua;Liu, Gang;Liu, Gang

作者机构：

关键词： CD1E; B2M; Cell-free protein synthesis; Protein expression; Membrane protein purification

期刊名称：PROTEIN EXPRESSION AND PURIFICATION （ 影响因子：1.6； 五年影响因子：1.6 ）

ISSN： 1046-5928

年卷期： 2023 年 203 卷

页码：

收录情况： SCI

摘要： CD1E, one of the most important glycolipid antigens on T cell membranes, is required for glycolipid antigen presentation on the cell surface. Cell-based recombinant expression systems have many limitations for synthe-sizing transmembrane proteins such as CD1E, including low protein yields and miss folding. To overcome these challenges, here we successfully synthesized high-quality soluble CD1E using an E.coli cell-free protein synthesis system (CFPS) with the aid of detergent. Following purification by Ni2+ affinity chromatography, we were able to obtain CD1E with >= 90% purity. Furthermore, we used the string website to predict the protein interaction network of CD1E and identified a potential binding partner B2M. Similarly, we synthesized soluble B2M in the E.coli CFPS. Finally, we verified the interaction between CD1E and B2M by using Surface Plasmon Resonance (SPR). Taken together, the methods described here provide an alternative way to obtain active transmembrane protein and may facilitate future structural and functional studies on CD1E.

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