Covalent modification of the renal Na+/H+ exchanger by N,N'-dicyclohexylcarbodiimide

P. Igarashi, P. S. Aronson

Research output: Contribution to journalArticle

37 Citations (Scopus)

Abstract

We studied the effect of the carboxyl group-specific reagent N,N'-dicyclohexylcarbodiimide on the Na+/H+ exchanger present in microvillus membrane vesicles isolated from rabbit renal cortices. Pretreatment of membrane vesicles with dicyclohexylcarbodiimide resulted in irreversible inhibition of Na+/H+ exchange which was not due to vesicle disruption or collapse of imposed pH gradients. Inhibition by dicyclohexylcarbodiimide followed pseudo-first-order kinetics, resulted primarily from a decrease in binding affinity for substrate, was pH-dependent in a manner consistent with reaction with carboxyl groups, and was greater than inhibition by hydrophilic carbodiimides. Substrates Na+ and Li+ and the competitive inhibitor amiloride protected against inhibition by dicyclohexylcarbodiimide in a pH-dependent fashion. Finally, we demonstrated amiloride-sensitive covalent binding of radiolabeled dicyclohexylcarbodiimide to a 100-kDa protein. In conclusion, a catalytically important carboxyl group is located in a relatively hydrophobic microenvironment at or near the external transport site of the renal Na+/H+ exchanger; and the transporter itself, or a subunit thereof, may be a 100-kDa protein.

Original languageEnglish (US)
Pages (from-to)860-868
Number of pages9
JournalJournal of Biological Chemistry
Volume262
Issue number2
StatePublished - 1987

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Dicyclohexylcarbodiimide
Sodium-Hydrogen Antiporter
Kidney
Amiloride
Membranes
Carbodiimides
Proton-Motive Force
Substrates
Microvilli
Proteins
Rabbits
Kinetics

ASJC Scopus subject areas

  • Biochemistry

Cite this

Covalent modification of the renal Na+/H+ exchanger by N,N'-dicyclohexylcarbodiimide. / Igarashi, P.; Aronson, P. S.

In: Journal of Biological Chemistry, Vol. 262, No. 2, 1987, p. 860-868.

Research output: Contribution to journalArticle

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