Effects of altering the Atp/Adp ratio on pump-mediated Na/K and Na/Na exchanges in resealed human red blood cell ghosts

Brian G. Kennedy, Gorm Lunn, Joseph F. Hoffman

Research output: Contribution to journalArticle

26 Scopus citations

Abstract

Resealed human red blood cell ghosts were prepared to contain a range of ADP concentrations at fixed ATP concentrations and vice versa. ATP/ADP ratios ranging from ~0.2 to 50 were set and maintained (for up to 45 min) in this system. ATP and ADP concentrations were controlled by the addition of either a phosphoarginine- or phosphocreatine-based regenerating system. Ouabain-sensitive unidirectional Na efflux was determined in the presence and absence of 15 mM external K as a function of the nucleotide composition. Na/K exchange was found to increase to saturation with ATP (K1/2 ≅ 250 µM), whereas Na/Na exchange (measured in K-free solutions) was a saturating function of ADP (K1/2 ≅ 350 µM). The elevation of ATP from ~100 to 1,800 jM did not appreciably affect Na/Na exchange. In the presence of external Na and a saturating concentration of external K, increasing the ADP concentration at constant ATP was found to decrease ouabain-sensitive Na/K exchange. The decreased Na/K exchange that still remained when the ADP/ATP ratio was high was stimulated by removal of external Na. Assuming that under normal substrate conditions the reaction cycle of the Na/K pump is rate-limited by the conformational change associated with the release of occluded K [E2. (K). ATP → E1 ATP + K], increasing ADP inhibits the rate of these transformations by competition with ATP for the E2(K) form. Aless likely alternative is that inhibition is due to competition with ATP at the high-affinity site (E1). The acceleration of the Na/K pump that occurs upon removing external Na at high levels of ADP evidently results from a shift in the forward direction of the transformation of the intermediates involved with the release of occluded Na from E1P. (Na). Thus, the nucleotide composition and the Na gradient can modulate the rate at which the Na/K. pump operates.

Original languageEnglish (US)
Pages (from-to)47-72
Number of pages26
JournalJournal of General Physiology
Volume87
Issue number1
DOIs
StatePublished - Jan 1 1986

ASJC Scopus subject areas

  • Physiology

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