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HCO 3 − and pH Modulate the Binding of Carbonic Anhydrase II to the Carboxy‐Terminus of kNBC1
Author(s) -
Bondar Galyna,
Pushkin Alexander,
Abuladze Natalia,
Kurtz Ira
Publication year - 2006
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.20.5.a1222-b
Subject(s) - carbonic anhydrase ii , chemistry , intracellular ph , cotransporter , carbonic anhydrase , bicarbonate , biophysics , pco2 , binding constant , binding site , biochemistry , intracellular , enzyme , medicine , sodium , biology , organic chemistry
We have previously reported that a complex of kNBC1 with carbonic anhydrase II (CAII) functions as a transport metabolon in vivo. Our studies indicated that interaction of CAII and kNBC1 is important for the function of the cotransporter and suggested that modulation of binding may significantly affect kNBC1‐mediated ion flux. In the carboxy‐terminus of kNBC1 (wt‐kNBC1‐ct), we previously identified two clusters of acidic amino acids involved in CAII binding suggesting that electrostatic interactions participate in this process. In this study we determined whether pH, Na + , K + , Cl − and HCO 3 − within their physiologic ranges altered the interaction between CAII and kNBC1‐ct. Both Na + and Cl − altered the binding between the two proteins with a maximum for both ions at ~ 20 mM whereas the effect of K + in the intracellular physiologic range (90–130 mM) was minimal. In non‐bicarbonate buffered solutions, the interaction of kNBC1‐ct and CAII was pH‐dependent with maximal binding at pH ~ 7.25. To determine the effect of changes in HCO 3 − /pCO 2 independent of pH changes, HCO 3 − was varied from 5–50 mM at a constant pH of 7.25 (by simultaneously varying the pCO 2 ). HCO 3 − /pCO 2 altered the binding of CAII and kNBC1‐ct at constant pH, with maximal binding at HCO 3 − /pCO 2 ~ 15 mM/40 mm Hg. The activity of CAII was unaffected by interaction with kNBC1‐ct. DIDS significantly decreased the activity of CAII (K i ~ 1.1 mM). In conclusion, the results demonstrate that the interaction of kNBC1‐ct and CAII is altered by the activity of various ions. DIDS not only inhibits kNBC1 but also independently decreases the activity of CAII. Regulation of the interaction of kNBC1‐ct with CAII by Na + , HCO 3 − and pH provides a potentially important mechanism for modulating the efficiency of intermolecular transfer of HCO 3 − between the two proteins. Supported by NIH.