Item

Abstract

Publisher Summary: This chapter discusses the structurally different nucleotide binding sites in Na, K-ATPase. Information on nucleotide binding sites of purified membrane-bound Na, K-ATPase isolated from the outer medulla of rat kidneys was obtained by three independent methods: (1) by measuring nucleotide effects on the inhibition of the enzyme with the ATP analog 6-[(3-carboxy-4-nitrophenyl)thiol]-98-V-ribo­furanosylpurine 5’-triphosphate (Nbs6ITP), (2) by measuring nucleotide effects on the enzyme inhibition with the SH-group reagent 5,5’-dithio­bis(2-nitrobenzoic acid) (DTNB), and (3) by analyzing the effects of ADP and AMP-PNP on the K⁺-dependent phosphatase activity of the Na, K-ATPase. The chapter graphically represents the inhibition of K⁺-dependent phosphatase activity by ADP in the presence and absence of additional nucleotides. The data presented in the chapter provide arguments for the existence of two simultaneously present nucleotide-binding sites in the Na, K-ATPase. The irreversible interaction of the Na, K­ATPase substrate Nbs6ITP is at nonhydrolyzing conditions differentially affected by low and high nucleotide concentrations. A nucleotide effect on the DTNB inactivation of the Na, K-ATPase observed in the absence of Na⁺ was specific for ATP and AMP-PNP but was not found with ADP.