Expression of functional human muscarinic M2 receptors in different insect cell lines
Expression of functional human muscarinic M2 receptors in different insect cell lines
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DOI：
10.3109/10799899709036611
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发表时间：
1997-01-01
期刊：
影响因子：
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通讯作者：
Guenet, C
中科院分区：
文献类型：
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作者：
Heitz, F;Nay, C;Guenet, C
使用杆状病毒表达系统在三种不同的昆虫细胞系Sf9、Sf21和High5中表达人M2受体。与Sf9细胞相比，Sf21细胞中的表达水平略微增加。相反，High5细胞不能产生比Sf9更多的重组蛋白。我们还表明，在这两个草地贪夜蛾细胞系的表达高峰后，感染6天达到，而在High5细胞，最大的表达发生在3天后。m2毒蕈碱受体的免疫检测清楚地表明，表达的蛋白质在Sf9和High5细胞中经历显著的蛋白水解，而在Sf21细胞中，这种现象是较难检测到的。此外，我们表明，在所有三个细胞系中，表达的重组受体的功能，因为它能够刺激GTP γ S结合在外源性G蛋白的存在下。提供了Sf 21和High5细胞中G蛋白（G α（i）、G α（o）和G β（常见））群体的分析。
Human M2 receptors were expressed using the baculovirus expression system in three different insect cell lines: Sf9, Sf21 and High5. The level of expression was slightly increased in Sf21 cells versus Sf9 cells. In contrast, High5 cells were not able to produce more recombinant protein than Sf9. We also show that in both Spodoptera frugiperda cell lines a peak of expression was reached after 6 days of infection, whereas in High5 cells, the maximum of expression occurred after 3 days. Immunodetection of m2 muscarinic receptor clearly shows that the expressed protein undergoes significant proteolysis in both the Sf9 and High5 cells, whereas in the Sf21 cells this phenomenon was less detectable. Additionally, we show that in all three cell lines, the expressed recombinant receptor was functional in that it was able to stimulate GTP gamma S binding in the presence of exogenous G-proteins. Analysis of the population of G-proteins (G alpha(i), G alpha(o) and G beta(common)) in Sf21 and High5 cells is provided.