人白细胞介素-13受体α2胞外域在毕赤酵母中的表达

Expression of human IL-13 receptor alpha2 extracellular domain in Pichia pastoris.

作者信息

Ohkuri Takatoshi, Takeda Chika, Yoshida Yuichiro, Izuhara Kenji, Imoto Taiji, Ueda Tadashi

机构信息

Department of Immunology, Graduate School of Pharmaceutical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan.

出版信息

Protein Expr Purif. 2007 Nov;56(1):48-53. doi: 10.1016/j.pep.2007.05.008. Epub 2007 May 27.

DOI:10.1016/j.pep.2007.05.008

PMID:17604644

Abstract

Interleukin-13 receptor alpha2 (IL-13Ralpha2) binds IL-13 with high affinity and plays an important role in IL-13 signaling as a decoy receptor. We expressed the extracellular domain of human IL-13Ralpha2 (1-313) in methylotrophic yeast Pichia pastoris. SDS-PAGE analysis by PAS staining and Western blot analysis detected the product of the extracellular domain of human IL-13Ralpha2 as glycoprotein from P. pastoris. The yield of purified extracellular domain of human IL-13Ralpha2 was 2mg from 1L of culture. From CD analysis, the 2D structure of the purified IL-13Ralpha2 showed the typical beta-sheet. ELISA of the purified IL-13Ralpha2 detected the binding activity for human IL-13. Thus, it was found that the active extracellular domain of human IL-13Ralpha2 was expressed from P. pastoris.

摘要

白细胞介素-13受体α2（IL-13Rα2）以高亲和力结合IL-13，并作为诱饵受体在IL-13信号传导中发挥重要作用。我们在甲基营养型酵母毕赤酵母中表达了人IL-13Rα2（1-313）的细胞外结构域。通过PAS染色的SDS-PAGE分析和蛋白质印迹分析检测到毕赤酵母中作为糖蛋白的人IL-13Rα2细胞外结构域的产物。从1升培养物中纯化的人IL-13Rα2细胞外结构域的产量为2毫克。通过圆二色性分析，纯化的IL-13Rα2的二维结构显示出典型的β-折叠。纯化的IL-13Rα2的酶联免疫吸附测定检测到对人IL-13的结合活性。因此，发现人IL-13Rα2的活性细胞外结构域在毕赤酵母中表达。