• Port-a-Patch

    世界上最小的膜片钳系统
  • Port-a-Patch

    易学易用——教学的理想工具
  • Port-a-Patch

    可记录细胞、细胞器与脂双层
  • Port-a-Patch

    世界上首款平板芯片膜片钳设备
  • Port-a-Patch

    可进行内灌流——更换细胞内液的理想工具

2014 - Membrane assembly of the functional KcsA potassium channel in a vesicle-based eukaryotic cell-free translation system

icon pap  Port-a-Patch and   icon vpp   Vesicle Prep Pro publication in Biosensors and Bioelectronics (2014)

Authors: 
Dondapati S.K., Kreir M., Quast R.B., Wüstenhagen D.A., Brüggemann A., Fertig N., Kubick S.

 

Journal: 
Biosens Bioelectron (2014) 15(59):174-83


Abstract: 

The potassium channel KcsA was heterologously expressed in a eukaryotic cell-free system. Both, the expression yields and functional analysis of the protein were reported. Qualitative and quantitative analyses of KcsA expression were performed by using 14C-labeled leucine as one of the amino acids supplemented in the cell-free reaction mixture. There was a time dependent increase in the protein yield as well as the intensity of the native tetramer band in insect cell derived microsomes. Electrophysiology measurements demonstrated the functional activity of the microsomes harboring KcsA showing single-channel currents with the typical biophysical characteristics of the ion channel. The channel behavior was asymmetric and showed positive rectification with larger currents towards positive voltages. KcsA channel currents were effectively blocked by potassium selective barium (Ba2+). This functional demonstration of an ion channel in eukaryotic cell-free system has a large potential for future applications including drug screening, diagnostic applications and functional assessment of complex membrane proteins like GPCRs by coupling them to ion channels in cell-free systems. Furthermore, membrane proteins can be expressed directly from linear DNA templates within 90 min, eliminating the need for additional cloning steps, which makes this cell-free system fast and efficient.


Download here

返回总览

We use cookies on our website. Some of them are essential for the operation of the site, while others help us to improve this site and the user experience (tracking cookies). You can decide for yourself whether you want to allow cookies or not. Please note that if you reject them, you may not be able to use all the functionalities of the site.