PERMEABILITY OF WILD-TYPE AND MUTANT CYSTIC-FIBROSIS TRANSMEMBRANE CONDUCTANCE REGULATOR CHLORIDE CHANNELS TO POLYATOMIC ANIONS

Permeability of the cystic fibrosis transmembrane conductance regulato r (CFTR) chloride channel to polyatomic anions of known dimensions was studied in stably transfected Chinese hamster ovary cells by using th e patch clamp technique. Biionic reversal potentials measured with ext ernal polyatomic anions gave the permeability ratio (P-X/P-Cl) sequenc e NO3- > Cl- > HCO3- > formate > acetate. The same selectivity sequenc e but somewhat higher permeability ratios were obtained when anions we re tested from the cytoplasmic side. Pyruvate, propanoate, methane sul fonate, ethane sulfonate, and gluconate were not measurably permeant ( P-X/P-Cl < 0.06) from either side of the membrane. The relationship be tween permeability ratios from the outside and ionic diameters suggest s a minimum functional pore diameter of similar to 5.3 Angstrom. Perme ability ratios also followed a lyotropic sequence, suggesting that per meability is dependent on ionic hydration energies. Site-directed muta genesis of two adjacent threonines in TM6 to smaller, less polar alani nes led to a significant (24%) increase in single channel conductance and elevated permeability to several large anions, suggesting that the se residues do not strongly bind permeating anions, but may contribute to the narrowest part of the pore.