Apparent affinity of CFTR for ATP is increased by continuous kinase activity.
The cystic fibrosis transmembrane conductance regulator (CFTR) is a chloride channel which is activated by protein phosphorylation and nucleoside triphosphates. We demonstrate here that fusion of the soluble catalytic subunit of cAMP-dependent protein kinase to the membrane protein bacteriorhodopsin yields a constitutively active protein kinase which activates CFTR effectively. As it is membrane-bound it is particularly useful for continuous perfusion of excised inside-out patches. We also tested the effect of a naturally membrane-bound protein kinase, cGMP-dependent protein kinase II, on CFTR. Both kinases, when continuously active, increase apparent affinity of CFTR to ATP about two-fold emphasizing the role of phosphorylation in modulating the interaction of ATP with the nucleotide binding domains.
Szellas
Tanjef
T
Max-Planck-Institut für Biophysik, Kennedyallee 70, D-60596 Frankfurt/Main, Germany.
Nagel
Georg
G
D000255:Adenosine Triphosphate / Q000378:metabolism* / Q000494:pharmacology; D000818:Animals; D001436:Bacteriorhodopsins / Q000235:genetics; D020134:Catalytic Domain / Q000235:genetics; D017868:Cyclic AMP-Dependent Protein Kinases / Q000235:genetics; D062967:Cyclic GMP-Dependent Protein Kinase Type II; D017869:Cyclic GMP-Dependent Protein Kinases / Q000235:genetics / Q000378:metabolism*; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics / Q000378:metabolism*; D004789:Enzyme Activation / Q000502:physiology; D006801:Humans; D009865:Oocytes / Q000378:metabolism; D018408:Patch-Clamp Techniques; D010766:Phosphorylation; D011485:Protein Binding / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism*; D014981:Xenopus
C505032:CFTR protein, human; D011993:Recombinant Fusion Proteins; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D001436:Bacteriorhodopsins; D000255:Adenosine Triphosphate; D017868:Cyclic AMP-Dependent Protein Kinases; D062967:Cyclic GMP-Dependent Protein Kinase Type II; D017869:Cyclic GMP-Dependent Protein Kinases; C569729:PRKG2 protein, human