title: Function of Shaker potassium channels produced by cell-free translation upon injection into Xenopus oocytes.
abstract: Voltage-gated ion channels are a class of membrane proteins that temporally orchestrate the ion flux critical for chemical and electrical signaling in excitable cells. Current methods to investigate the function of these channels rely on heterologous expression in living systems or reconstitution into artificial membranes; however these approaches have inherent drawbacks which limit potential biophysical applications. Here, we describe a new integrated approach combining cell-free translation of membrane proteins and in vivo expression using Xenopus laevis oocytes. In this method, proteoliposomes containing Shaker potassium channels are synthesized in vitro and injected into the oocytes, yielding functional preparations as shown by electrophysiological and fluorescence measurements within few hours. This strategy for studying eukaryotic ion channels is contrasted with existing, laborious procedures that require membrane protein extraction and reconstitution into synthetic lipid systems.
authors.affiliation: Department of Neuroscience, University of Wisconsin-Madison, Madison, WI 53706, USA.
authors.forename: Brian W
authors.initials: BW
authors.lastname: Jarecki
authors.forename: Shin-ichi
authors.initials: S
authors.lastname: Makino
authors.forename: Emily T
authors.initials: ET
authors.lastname: Beebe
authors.forename: Brian G
authors.initials: BG
authors.lastname: Fox
authors.forename: Baron
authors.initials: B
authors.lastname: Chanda
mesh_terms: D000818:Animals; D002474:Cell-Free System; D008564:Membrane Potentials / Q000502:physiology; D008565:Membrane Proteins / Q000096:biosynthesis / Q000378:metabolism; D009865:Oocytes / Q000166:cytology* / Q000378:metabolism*; D011510:Proteolipids / Q000096:biosynthesis / Q000378:metabolism; D051637:Shaker Superfamily of Potassium Channels / Q000378:metabolism* / Q000502:physiology; D014982:Xenopus laevis / Q000502:physiology*
chemical_list: D008565:Membrane Proteins; D011510:Proteolipids; D051637:Shaker Superfamily of Potassium Channels; C020485:proteoliposomes