Related Experiment Video
Updated: May 27, 2026

Reconstitution of a Kv Channel into Lipid Membranes for Structural and Functional Studies
Published on: July 13, 2013
Reconstitution of KCNE1 into lipid bilayers: comparing the structural, dynamic, and activity differences in micelle
Aaron T Coey1, Indra D Sahu, Thusitha S Gunasekera
1Department of Chemistry and Biochemistry, Miami University, Oxford, Ohio 45056, United States.
Abstract:
KCNE1 (minK), found in the human heart and cochlea, is a transmembrane protein that modulates the voltage-gated potassium KCNQ1 channel. While KCNE1 has previously been the subject of extensive structural studies in lyso-phospholipid detergent micelles, key observations have yet to be confirmed and refined in lipid bilayers. In this study, a reliable method for reconstituting KCNE1 into lipid bilayer vesicles composed of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) and 1-palmitoyl-2-oleoyl-sn-glycero-3-phospho(1'-rac-glycerol) (sodium salt) (POPG) was developed. Microinjection of the proteoliposomes into Xenopus oocytes expressing the human KCNQ1 (K(V)7.1) voltage-gated potassium channel led to nativelike modulation of the channel. Circular dichroism spectroscopy demonstrated that the percent helicity of KCNE1 is significantly higher for the protein reconstituted in lipid vesicles than for the previously described structure in 1.0% 1-myristoyl-2-hydroxy-sn-glycero-3-phospho(1'-rac-glycerol) (sodium salt) (LMPG) micelles. SDSL electron paramagnetic resonance spectroscopic techniques were used to probe the local structure and environment of Ser28, Phe54, Phe57, Leu59, and Ser64 of KCNE1 in both POPC/POPG vesicles and LMPG micelles. Spin-labeled KCNE1 cysteine mutants at Phe54, Phe57, Leu59, and Ser64 were found to be located inside POPC/POPG vesicles, whereas Ser28 was found to be located outside the membrane. Ser64 was shown to be water inaccessible in vesicles but found to be water accessible in LMPG micelle solutions. These results suggest that key components of the micelle-derived structure of KCNE1 extend to the structure of this protein in lipid bilayers but also demonstrate the need to refine this structure using data derived from the bilayer-reconstituted protein to more accurately define its native structure. This work establishes the basis for such future studies.
More Related Videos
11:42Reconstitution of a Transmembrane Protein, the Voltage-gated Ion Channel, KvAP, into Giant Unilamellar Vesicles for Microscopy and Patch Clamp Studies
Published on: January 22, 2015
10:19In Vitro Reconstitution of the Actin Cytoskeleton Inside Giant Unilamellar Vesicles
Published on: August 25, 2022
Related Concept Videos
Pinching-off of Coated Vesicles
Membrane Fluidity
Mosaic nature of the membrane
The mosaic characteristic of the membrane helps the plasma membrane remain fluid. The integral proteins and lipids exist as separate but loosely-attached molecules in the membrane. The membrane is a relatively...
Assembly of the Lipid Bilayer in the ER
A large chunk of any biological membrane is composed of phospholipids. These lipids have a heterogeneous distribution across different subcellular organelles and even between...
COP Coated Vesicles
Asymmetric Lipid Bilayer