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Two‐Electrode Voltage‐Clamp Analysis of Na,K‐ATPase Asparagine 776 Mutants
Author(s) -
KOENDERINK JAN B.,
GEIBEL SVEN,
GRABSCH EVA,
PONT JAN JOEP H. H. M.,
BAMBERG ERNST,
FRIEDRICH THOMAS
Publication year - 2003
Publication title -
annals of the new york academy of sciences
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.712
H-Index - 248
eISSN - 1749-6632
pISSN - 0077-8923
DOI - 10.1111/j.1749-6632.2003.tb07152.x
Subject(s) - chemistry , extracellular , membrane potential , mutant , biophysics , depolarization , steady state (chemistry) , wild type , asparagine , membrane , voltage clamp , enzyme , stereochemistry , biochemistry , biology , gene
A bstract : Steady‐state and pre‐steady‐state currents of Asn 776 mutants of Na,K‐ATPase are presented. The stationary current generated by N776Q strongly depends on the membrane potential, but has a negative slope, opposite to that of the wild‐type enzyme. The apparent rate constant of the reaction sequence E 1 P(Na + ) ↔ E 2 P + Na + of this mutant is rather independent of the membrane potential and is at resting and depolarizing membrane potential higher than that of the wild‐type enzyme. Thus, the voltage‐dependent increase of the rate coefficient of the wild type that is associated with extracellular Na + rebinding is almost absent in the N776Q mutant. These findings indicate that dislocating the carboxamide group of Asn 776 decreases the affinity of sodium at its extracellular binding site.