Purification of the functional plant membrane channel KAT1.
The inward-rectifying K(+) channel KAT1 is expressed mainly in Arabidopsis thaliana guard cells. The purification of functional KAT1 has never been reported. We investigated the extraction of the plant K(+) channel KAT1 with different detergents, as an example for how to select detergents for purifying a eukaryotic membrane protein. A KAT1-GFP fusion protein was used to screen a library of 46 detergents for the effective solubilization of intact KAT1. Then, a "test set" of three detergents was picked for further analysis, based on their biochemical characteristics and availability. The combination use of the selected detergents enabled the effective purification of functional KAT1 with affinity and gel-filtration chromatography.
Hibi
Takao
T
Department of Bioscience, Fukui Prefectural University, 4-1-1 MatsuokaKenjyojima, Eiheiji-cho, Fukui 910-1195, Japan. hibi@fpu.ac.jp
Aoki
Shiho
S
Oda
Keisuke
K
Munemasa
Shintaro
S
Ozaki
Shunsuke
S
Shirai
Osamu
O
Murata
Yoshiyuki
Y
Uozumi
Nobuyuki
N
D000818:Animals; D029681:Arabidopsis Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*; D002846:Chromatography, Affinity / Q000379:methods; D002850:Chromatography, Gel; D003902:Detergents / Q000737:chemistry*; D005453:Fluorescence; D049452:Green Fluorescent Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D006639:Histidine / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D007313:Insecta / Q000166:cytology; D024661:Potassium Channels, Inwardly Rectifying / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*
D029681:Arabidopsis Proteins; D003902:Detergents; C501210:KAT1 protein, Arabidopsis; D024661:Potassium Channels, Inwardly Rectifying; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D006639:Histidine