Sharapova Olga A, Pozdnykova Natalia V, Laurinavichyute Daniela K, Yurkova Maria S, Posypanova Galina A, Fedorov Alexey N, Severin Sergey E, Severin Evgeny S
Moscow Research Institute of Medical Ecology, Simpheropolski Blvd. 8, Moscow 117638, Russia.
Protein Expr Purif. 2010 Sep;73(1):31-5. doi: 10.1016/j.pep.2010.03.025. Epub 2010 Apr 2.
Human alpha-fetoprotein (hAFP) is an oncofetal protein which is a common cancer marker. Conjugates of native hAFP with different cytostatic agents inhibit growth of cancer cells in vivo and in vitro. The hAFP interacts with its receptor (AFPR) on the surface of cancer cells via its C-terminal domain. The aim of this work was to develop a highly efficient expression system in Escherichia coli and efficient refolding procedure for the recombinant C-terminal fragment of hAFP (rAFP-Cterm) and to characterize its functional properties. C-terminal fragment of hAFP (rAFP-Cterm) comprising amino acids from 404 to 609 was expressed in E. coli BL21 (DE3) strain with high yield. High efficient purification and refolding procedures were developed giving yield of refolded protein about 80% with purity about 95%. The refolded rAFP-Cterm bound specifically with cancer cells carrying AFPR and was accumulated by them with the same efficiency as native hAFP. This rAFP-Cterm can be used as a vehicle for the targeted delivery of drugs to cancer cells.
人甲胎蛋白（hAFP）是一种癌胚蛋白，是一种常见的癌症标志物。天然hAFP与不同细胞抑制剂的缀合物在体内和体外均可抑制癌细胞生长。hAFP通过其C末端结构域与癌细胞表面的受体（AFPR）相互作用。本研究的目的是在大肠杆菌中开发一种高效表达系统，并为重组hAFP C末端片段（rAFP-Cterm）建立高效的复性方法，同时对其功能特性进行表征。包含404至609位氨基酸的hAFP C末端片段（rAFP-Cterm）在大肠杆菌BL21（DE3）菌株中高产表达。开发了高效的纯化和复性方法，复性蛋白的产率约为80%，纯度约为95%。复性后的rAFP-Cterm与携带AFPR的癌细胞特异性结合，并以与天然hAFP相同的效率被它们摄取。这种rAFP-Cterm可作为将药物靶向递送至癌细胞的载体。
