TY - JOUR
T1 - Heterologous expression and purification of an active human TRPV3 ion channel
AU - Kol, S.
AU - Braun, Christian Seeberg
AU - Thiel, G.
AU - Doyle, D.A.
AU - Sundström, M.
AU - Gourdon, Pontus Emanuel
AU - Nissen, Poul
PY - 2013/12/1
Y1 - 2013/12/1
N2 - The transient receptor potential vanilloid 3 (TRPV3) cation channel is widely expressed in human tissues and has been shown to be activated by mild temperatures or chemical ligands. In spite of great progress in the TRP-channel characterization, very little is known about their structure and interactions with other proteins at the atomic level. This is mainly caused by difficulties in obtaining functionally active samples of high homogeneity. Here, we report on the high-level Escherichia coli expression of the human TRPV3 channel, for which no structural information has been reported to date. We selected a suitable detergent and buffer system using analytical size-exclusion chromatography and a thermal stability assay. We demonstrate that the recombinant purified protein contains high α-helical content and migrates as dimers and tetramers on native PAGE. Furthermore, the purified channel also retains its current inducing activity, as shown by electrophysiology experiments. The ability to produce the TRPV3 channel heterologously will aid future functional and structural studies.
AB - The transient receptor potential vanilloid 3 (TRPV3) cation channel is widely expressed in human tissues and has been shown to be activated by mild temperatures or chemical ligands. In spite of great progress in the TRP-channel characterization, very little is known about their structure and interactions with other proteins at the atomic level. This is mainly caused by difficulties in obtaining functionally active samples of high homogeneity. Here, we report on the high-level Escherichia coli expression of the human TRPV3 channel, for which no structural information has been reported to date. We selected a suitable detergent and buffer system using analytical size-exclusion chromatography and a thermal stability assay. We demonstrate that the recombinant purified protein contains high α-helical content and migrates as dimers and tetramers on native PAGE. Furthermore, the purified channel also retains its current inducing activity, as shown by electrophysiology experiments. The ability to produce the TRPV3 channel heterologously will aid future functional and structural studies.
UR - http://www.scopus.com/inward/record.url?scp=84888305651&partnerID=8YFLogxK
U2 - 10.1111/febs.12520
DO - 10.1111/febs.12520
M3 - Journal article
C2 - 24028292
AN - SCOPUS:84888305651
SN - 1742-4658
VL - 280
SP - 6010
EP - 6021
JO - The F E B S Journal (Online)
JF - The F E B S Journal (Online)
IS - 23
ER -