Muscarinic acetylcholine receptor produced in recombinant baculovirus infected Sf9 insect cells couples with endogenous G‐proteins to activate ion channels

FEBS Letters

Vasudevan, Subhash⁴  Chung, Shin-Ho¹  Gage, Peter W.²  Premkumar, Louis²  Reiländer, Helmut³  Stowe, Sally⁵ 

[1] Protein Dynamics Unit, Department of Chemistry, Australian National University, Canberra, ACT 2601, Australia;John Curtin School of Medical Research, Australian National University, Canberra, ACT 2601, Australia;Max Planck Institut für Biophysik, Frankfurt, Germany;Research School of Chemistry, Australian National University, Canberra, ACT 2601, Australia;Research School of Biological Sciences, Australian National University, Canberra, ACT 2601, Australia

关键词: Muscarinic acetylcholine receptor;    Baculovirus expression;    Patch clamp;    G-protein;    Potassium channel;   

DOI  :  10.1016/0014-5793(92)81354-O

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

Following the infection of insect ovarian cells (Sf9) with recombinant baculovirus bearing the cDNA coding for the rat muscarinic acetylcholine (ACh) receptor subtype m3, ionic flux across the membrane in response to the application of ACh was examined electrophysiologically. We show that ACh activates potassium currents. The response is abolished when cells are treated with pertussis toxin. No ACh-induced currents are observed from uninfected cells or cells infected with virus which do not contain the cDNA coding for ACh receptors in its genome. The characteristics of single channel currents show time-dependent changes following the application of ACh. Initially, ACh activates brief channel currents with a conductance of about 5 pS. The conductance level of channels gradually increases in steps to 10 pS and then to 20 pS and 40 pS. At the same time, channel open probability also increases. Thereafter, additional channels appear, opening and closing independently of, or at times in synchrony with, the original channel.

【 授权许可】

Unknown