Binding of 2′(3′)-O-(2,4,6-Trinitrophenyl)ADP to Soluble αβ Protomers of Na,K-ATPase Modified with Fluorescein Isothiocyanate
Open Access
- 1 May 1996
- journal article
- Published by Elsevier BV
- Vol. 271 (21), 12317-12321
- https://doi.org/10.1074/jbc.271.21.12317
Abstract
The overall reaction of well-defined solubilized protomers of Na,K-ATPase (one alpha plus one beta subunit) retains the dual ATP dependence observed with the membrane-bound enzyme, with distinctive ATP effects in the submicromolar and submillimolar ranges (Ward, D. G., and Cavieres, J. D. (1993) Proc. Natl. Acad. Sci. U. S. A. 90, 5332-5336). We have now found that the K+/-phosphatase activity of the alpha beta protomers is still inhibited by 2'(3')-O-(2,4,6-trinitrophenyl)adenosine 5'-diphosphate (TNP-ADP). What is most significant is that the TNP-ADP effect can be observed clearly with protomeric enzyme whose high affinity ATP site has been blocked covalently with fluorescein isothiocyanate. We conclude that nucleotides can bind at two discrete sites in each protomeric unit of Na,K-ATPase.Keywords
This publication has 29 references indexed in Scilit:
- Identification of an Amino Acid in the ATP Binding Site of Na+/K+-ATPase after Photochemical Labeling with 8-Azido-ATPBiochemistry, 1994
- Simultaneous binding of phosphate and TNP-ADP to FITC-modified sodium-potassium-ATPaseBiochemistry, 1993
- Phosphate binding and ATP‐binding sites coexist in Na+/K+‐transporting ATPase, as demonstrated by the inactivating MgPO4 complex analogue Co(NH3)4PO4European Journal of Biochemistry, 1991
- Substrate sites of the (Na+ + K+)-ATPase: Pertinence of the adenine and fluorescein binding sitesBiochimica et Biophysica Acta (BBA) - Protein Structure and Molecular Enzymology, 1988
- The molecular size required varies according to the reaction step round the sodium pump cycleFEBS Letters, 1987
- Affinity modification of E1‐form of Na+, K+ ‐ATPase revealed Asp‐710 in the catalytic siteFEBS Letters, 1987
- Inhibition of ion pump ATPase activity by 3′-O-(4-Benzoyl)benzoyl-ATP (BzATP): assessment of BzATP as an active site-directed probeBiochimica et Biophysica Acta (BBA) - Biomembranes, 1986
- Evidence for a diprotomeric structure of Na,K‐ATPaseFEBS Letters, 1986
- The amino acid sequence of the fluorescein isothiocyanate reactive site of lamb and rat kidney Na+- and K+-dependent ATPaseBiochemical and Biophysical Research Communications, 1984
- Purification and characterization of (Na+, K+)-ATPase. V. Conformational changes in the enzyme. Transitions between the Na-form and the K-form studied with tryptic digestion as a toolBiochimica et Biophysica Acta (BBA) - Biomembranes, 1975