Biochemical Activity of NHX2.
(A) Complementation of the yeast nhx1 mutant (top), transformed with an empty vector, or to express NHX1, NHX2, or His-tagged NHX2 (four independent transformants), in Arg phosphate medium supplemented with 70 mM NaCl and yeast peptone dextrose (YPD) medium with 30 μg/mL hygromycin B (HygB).
(B) Typical exchange reaction in proteoliposomes. Arrows indicate the addition of substrate salts NaCl or KCl (1). SO4(NH4)2 was added to collapse the proton gradient and end the assay (2). Pyranine fluorescence at 510 nm is given in arbitrary units.
(C) Saturation kinetics of exchange rates catalyzed by NHX2 as a function of substrate (NaCl) concentration.
(D) Exchange rates using as substrate 45 mM sodium (NaGlu) or K+ (KGlu) gluconate salts, 22.5 mM each of these salts (Na/K), and 45 mM NaCl or KCl. Shown is the mean exchange rates of two independent assays.