Methods to study CFTR protein in vitro.
CFTR is a cyclic AMP and nucleotide-related chloride-selective channel with a low unitary conductance. Many of the physiological roles of CFTR are effectively studied in intact cells and tissues. However, there are also several clear advantages to the application of cell-free technologies to the study of the biochemical and biophysical properties of CFTR. When expressed in heterologous cells, CFTR is processed relatively poorly, depending, however, on the cell-type analysed. In some cells, only 20-25% of the protein which is initially synthesized exits the endoplasmic reticulum to insert into the cell membrane [Cell 83 (1995) 121; EMBO J. 13 (1994) 6076]. Further, many of the disease-causing mutants of CFTR result in even lower processing efficiencies. Therefore, several procedures have been developed to study regulated CFTR channel function expressed in microsomal membranes and following its purification and reconstitution. These experimental approaches and their application are discussed here.
Department of Physiology and Biophysics, University of Alabama at Birmingham, Birmingham, AL, USA.
Dale J
DJ
Benos
Bakhrom K
BK
Berdiev
Iskander I
II
Ismailov
Lynda S
LS
Ostedgaard
Ilana
I
Kogan
Canhui
C
Li
Mohabir
M
Ramjeesingh
Christine E
CE
Bear
D002462:Cell Membrane / Q000502:physiology*; D019411:Clinical Laboratory Techniques; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000096:biosynthesis* / Q000235:genetics / Q000502:physiology; D006801:Humans; D066298:In Vitro Techniques; D008565:Membrane Proteins / Q000096:biosynthesis / Q000235:genetics / Q000502:physiology
C505032:CFTR protein, human; D008565:Membrane Proteins; D019005:Cystic Fibrosis Transmembrane Conductance Regulator