Bacterial synthesis, purification, and solubilization of membrane protein KCNE3, a regulator of voltage-gated potassium channels

An efficient method is described for production of membrane protein KCNE3 and its isotope labeled derivatives (¹⁵N-, ¹⁵N-/¹³C-) in amounts sufficient for structural-functional investigations. The purified protein preparation within different detergent micelles was characterized using dynamic light scattering, CD spectroscopy, and NMR spectroscopy. It is shown that within DPC/LDAO micelles the protein is in monomeric form and acquires mainly α-helical conformation. The existence of cross-peaks for all glycines of the ¹⁵N-HSQC NMR spectra as well as relatively small line widths (∼20 Hz) confirm the high quality of the preparation and the possibility of obtaining structural-dynamic information on KCNE3 by high resolution heteronuclear NMR spectroscopy.