Document Detail

Effect of human carbonic anhydrase II on the activity of the human electrogenic Na/HCO3 cotransporter NBCe1-A in Xenopus oocytes.
MedLine Citation:
PMID:  16687407     Owner:  NLM     Status:  MEDLINE    
Others report that carbonic anhydrase II (CA II) binds to the C termini of the anion exchanger AE1 and the electrogenic Na/HCO3 cotransporter NBCe1-A, enhancing transport. After injecting oocytes with NBCe1-A cRNA (Day 0), we measured NBC current (I(NBC)) by two-electrode voltage clamp (Day 3), injected CA II protein + Tris or just Tris (Day 3), measured I(NBC) or the initial rate at which the intracellular pH fell (dpH(i)/dt) upon applying 5% CO2 (Day 4), exposed oocytes to the permeant CA inhibitor ethoxzolamide (EZA), and measured I(NBC) or dpH(i)/dt (Day 4). Because dpH(i)/dt was greater in CA II than Tris oocytes, and EZA eliminated the difference, injected CA II was functional. I(NBC) slope conductance was unaffected by injecting CA II. Moreover, EZA had identical effects in CA II versus Tris oocytes. Thus, injected CA II does not enhance NBC activity. In a second protocol, we made a fusion protein with enhanced green fluorescent protein (EGFP) at the 5' end of NBCe1-A and CA II at the 3' end (EGFP-e1-CAII). We measured I(NBC) or dpH(i)/dt (days 3-4), exposed oocytes to EZA, and measured I(NBC) or dpH(i)/dt (Day 3-4). dpH(i)/dt was greater in oocytes expressing EGFP-e1-CA II versus EGFP-e1, and EZA eliminated the difference. Thus, fused CA II was functional. Slope conductances of EGFP-e1-CAII versus EGFP-e1 oocytes were indistinguishable, and EZA had no effect. Thus, even when fused to NBCe1-A, CA II does not enhance NBCe1-A activity.
Jing Lu; Christopher M Daly; Mark D Parker; Harindarpal S Gill; Peter M Piermarini; Marc F Pelletier; Walter F Boron
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Publication Detail:
Type:  Journal Article; Research Support, N.I.H., Extramural; Research Support, Non-U.S. Gov't     Date:  2006-05-10
Journal Detail:
Title:  The Journal of biological chemistry     Volume:  281     ISSN:  0021-9258     ISO Abbreviation:  J. Biol. Chem.     Publication Date:  2006 Jul 
Date Detail:
Created Date:  2006-07-10     Completed Date:  2006-08-21     Revised Date:  2014-09-05    
Medline Journal Info:
Nlm Unique ID:  2985121R     Medline TA:  J Biol Chem     Country:  United States    
Other Details:
Languages:  eng     Pagination:  19241-50     Citation Subset:  IM    
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MeSH Terms
Carbonic Anhydrase II / chemistry*
Ethoxzolamide / pharmacology
Green Fluorescent Proteins / metabolism
Hydrogen-Ion Concentration
Models, Biological
Oocytes / metabolism*
RNA, Complementary / metabolism
Sodium-Bicarbonate Symporters / metabolism*,  physiology
Time Factors
Xenopus / metabolism*
Grant Support
Reg. No./Substance:
0/RNA, Complementary; 0/SLC4A4 protein, human; 0/Sodium-Bicarbonate Symporters; 147336-22-9/Green Fluorescent Proteins; EC 4.2.1.-/Carbonic Anhydrase II; Z52H4811WX/Ethoxzolamide

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