Carbonic anhydrase II increases the activity of the human electrogenic Na+/HCO3- cotransporter.
Several acid/base-coupled membrane transporters, such as the electrogenic sodium-bicarbonate cotransporter (NBCe1), have been shown to bind to different carbonic anhydrase isoforms to create a "transport metabolon." We have expressed NBCe1 derived from human kidney in oocytes of Xenopus leavis and determined its transport activity by recording the membrane current in voltage clamp, and the cytosolic H(+) and Na(+) concentrations using ion-selective microelectrodes. When carbonic anhydrase isoform II (CAII) had been injected into oocytes, the membrane current and the rate of cytosolic Na(+) rise, indicative for NBCe1 activity, increased significantly with the amount of injected CAII (2-200 ng). The CAII inhibitor ethoxyzolamide reversed the effects of CAII on the NBCe1 activity. Co-expressing wild-type CAII or NH(2)-terminal mutant CAII together with NBCe1 provided similar results, whereas co-expressing the catalytically inactive CAII mutant V143Y had no effect on NBCe1 activity. Mass spectrometric analysis and the rate of cytosolic H(+) change following addition of CO(2)/HCO(3)(-) confirmed the catalytic activity of injected and expressed CAII in oocytes. Our results show that the transport capacity of NBCe1 is enhanced by the catalytic activity of CAII, in line with the notion that CAII forms a transport metabolon with NBCe1.
Becker
Holger M
HM
Abteilung für Allgemeine Zoologie, Fachbereich Biologie, Technische Universität Kaiserslautern, P. O. Box 3049, D-67653 Kaiserslautern, Germany. h.becker@biologie.uni-kl.de
Deitmer
Joachim W
JW
D019943:Amino Acid Substitution; D000818:Animals; D024402:Carbonic Anhydrase II / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D002257:Carbonic Anhydrase Inhibitors / Q000494:pharmacology; D005016:Ethoxzolamide / Q000494:pharmacology; D015870:Gene Expression; D006801:Humans; D006859:Hydrogen / Q000378:metabolism; D017136:Ion Transport / Q000187:drug effects / Q000235:genetics; D007668:Kidney / Q000378:metabolism; D008564:Membrane Potentials / Q000187:drug effects / Q000235:genetics; D008839:Microelectrodes; D020125:Mutation, Missense; D009865:Oocytes / Q000166:cytology; D012964:Sodium / Q000378:metabolism; D027982:Sodium-Bicarbonate Symporters / Q000235:genetics / Q000378:metabolism*; D014981:Xenopus
D002257:Carbonic Anhydrase Inhibitors; D027982:Sodium-Bicarbonate Symporters; D006859:Hydrogen; D012964:Sodium; D024402:Carbonic Anhydrase II; D005016:Ethoxzolamide