HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Eakle, K. A. Articles by Farley, R. A. Search for Related Content PubMed PubMed Citation Articles by Eakle, K. A. Articles by Farley, R. A. Social Bookmarking                      What's this?

Volume 270, Number 23, Issue of June 9, pp. 13937-13947, 1995
©1995 by The American Society for Biochemistry and Molecular Biology, Inc.

---

The Influence of Subunit Structure on the Interaction of Na/K-ATPase Complexes with Na
A CHIMERIC SUBUNIT REDUCES THE Na DEPENDENCE OF PHOSPHOENZYME FORMATION FROM ATP

Kurt A. Eakle , Rong-Ming Lyu , Robert A. Farley

High-affinity ouabain binding to Na/K-ATPase (sodium- and potassium-transport adenosine triphosphatase (EC 3.6.1.37)) requires phosphorylation of the subunit of the enzyme either by ATP or by inorganic phosphate. For the native enzyme (/1), the ATP-dependent reaction proceeds about 4-fold more slowly in the absence of Na than when saturating concentrations of Na are present. Hybrid pumps were formed from either the 1 or the 3 subunit isoforms of Na/K-ATPase and a chimeric subunit containing the transmembrane segment of the Na/K-ATPase 1 isoform and the external domain of the gastric H/K-ATPase subunit (/NH1 complexes). In the absence of Na, these complexes show a rate of ATP-dependent ouabain binding from 75-100% of the rate seen in the presence of Na depending on buffer conditions. Nonhydrolyzable nucleotides or treatment of ATP with apyrase abolishes ouabain binding, demonstrating that ouabain binding to /NH1 complexes requires phosphorylation of the protein. Buffer ions inhibit ouabain binding by /NH1 in the absence of Na rather than promote ouabain binding, indicating that they are not substituting for sodium ions in the phosphorylation reaction. The pH dependence of ATP-dependent ouabain binding in the presence or absence of Na is similar, suggesting that protons are probably not substituting for Na. Hybrid /NH1 pumps also show slightly higher apparent affinities (2-3-fold) for ATP, Na, and ouabain; however, these are not sufficient to account for the increase in ouabain binding in the absence of Na. In contrast to phosphoenzyme formation and ouabain binding by /NH1 complexes in the absence of Na, ATPase activity, measured as release of phosphate from ATP, requires Na. These data suggest that the transition from EP to EP during the catalytic cycle does not occur when the sodium binding sites are not occupied. Thus, the chimeric subunit reduces or eliminates the role of Na in phosphoenzyme formation from ATP, but Na binding or release by the enzyme is still required for ATP hydrolysis and release of phosphate.

CiteULike

Complore

Connotea

Del.icio.us

Digg

Reddit

Technorati

This article has been cited by other articles:

				I. Dostanic, J. N. Lorenz, J. E. J. Schultz, I. L. Grupp, J. C. Neumann, M. A. Wani, and J. B Lingrel The {alpha}2 Isoform of Na,K-ATPase Mediates Ouabain-induced Cardiac Inotropy in Mice J. Biol. Chem., December 26, 2003; 278(52): 53026 - 53034. [Abstract] [Full Text] [PDF]

				B. Huang, G. Blanco, R. W. Mercer, T. Fleming, and J. S. Pepose Human Corneal Endothelial Cell Expression of Na+,K+-Adenosine Triphosphatase Isoforms Arch Ophthalmol, June 1, 2003; 121(6): 840 - 845. [Abstract] [Full Text] [PDF]

				G. Sweeney, W. Niu, V. A. Canfield, R. Levenson, and A. Klip Insulin increases plasma membrane content and reduces phosphorylation of Na+-K+ pump alpha 1-subunit in HEK-293 cells Am J Physiol Cell Physiol, December 1, 2001; 281(6): C1797 - C1803. [Abstract] [Full Text] [PDF]

				S. He, D. A. Shelly, A. E. Moseley, P. F. James, J. H. James, R. J. Paul, and J. B. Lingrel The {alpha}1- and {alpha}2-isoforms of Na-K-ATPase play different roles in skeletal muscle contractility Am J Physiol Regulatory Integrative Comp Physiol, September 1, 2001; 281(3): R917 - R925. [Abstract] [Full Text] [PDF]

				A. Shainskaya, A. Schneeberger, H.-J. Apell, and S. J. D. Karlish Entrance Port for Na+ and K+ Ions on Na+,K+-ATPase in the Cytoplasmic Loop between Trans-membrane Segments M6 and M7 of the alpha Subunit. PROXIMITY OF THE CYTOPLASMIC SEGMENT OF THE beta SUBUNIT J. Biol. Chem., January 21, 2000; 275(3): 2019 - 2028. [Abstract] [Full Text] [PDF]

				C. R. Burrow, O. Devuyst, X. Li, L. Gatti, and P. D. Wilson Expression of the beta 2-subunit and apical localization of Na+-K+-ATPase in metanephric kidney Am J Physiol Renal Physiol, September 1, 1999; 277(3): F391 - F403. [Abstract] [Full Text] [PDF]

				S. Asano, T. Kimura, S. Ueno, M. Kawamura, and N. Takeguchi Chimeric Domain Analysis of the Compatibility between H+,K+-ATPase and Na+,K+-ATPase beta -Subunits for the Functional Expression of Gastric H+,K+-ATPase J. Biol. Chem., August 6, 1999; 274(32): 22257 - 22265. [Abstract] [Full Text] [PDF]

				F. Jaisser and A. T. Beggah The nongastric H+-K+-ATPases: molecular and functional properties Am J Physiol Renal Physiol, June 1, 1999; 276(6): F812 - F824. [Abstract] [Full Text] [PDF]

				J. B. Koenderink, H. G. P. Swarts, H. P. H. Hermsen, and J. J. H. H. M. De Pont The {beta}-Subunits of Na+,K+-ATPase and Gastric H+,K+-ATPase Have a High Preference for Their Own {alpha}-Subunit and Affect the K+ Affinity of These Enzymes J. Biol. Chem., April 23, 1999; 274(17): 11604 - 11610. [Abstract] [Full Text] [PDF]

				U. Hasler, X. Wang, G. Crambert, P. Beguin, F. Jaisser, J.-D. Horisberger, and K. Geering Role of beta -Subunit Domains in the Assembly, Stable Expression, Intracellular Routing, and Functional Properties of Na,K-ATPase J. Biol. Chem., November 13, 1998; 273(46): 30826 - 30835. [Abstract] [Full Text] [PDF]

				S.-G. Wang and R. A. Farley Valine 904, Tyrosine 898, and Cysteine 908 in Na,K-ATPase alpha  Subunits Are Important for Assembly with beta  Subunits J. Biol. Chem., November 6, 1998; 273(45): 29400 - 29405. [Abstract] [Full Text] [PDF]

				G. Blanco and R. W. Mercer Isozymes of the Na-K-ATPase: heterogeneity in structure, diversity in function Am J Physiol Renal Physiol, November 1, 1998; 275(5): F633 - F650. [Abstract] [Full Text] [PDF]

				E. Arystarkhova and K. J. Sweadner Tissue-specific Expression of the Na,K-ATPase beta 3 Subunit. THE PRESENCE OF beta 3 IN LUNG AND LIVER ADDRESSES THE PROBLEM OF THE MISSING SUBUNIT J. Biol. Chem., September 5, 1997; 272(36): 22405 - 22408. [Abstract] [Full Text] [PDF]

				J. Codina, B. C. Kone, J. T. Delmas-Mata, and T. D. DuBose Jr. Functional Expression of the Colonic H+,K+-ATPase alpha -Subunit. PHARMACOLOGIC PROPERTIES AND ASSEMBLY WITH X+,K+-ATPase beta -SUBUNITS J. Biol. Chem., November 22, 1996; 271(47): 29759 - 29763. [Abstract] [Full Text] [PDF]

				B. Fiedler and G. Scheiner-Bobis Transmembrane Topology of alpha - and beta -Subunits of Na+,K+-ATPase Derived from beta -Galactosidase Fusion Proteins Expressed in Yeast J. Biol. Chem., November 15, 1996; 271(46): 29312 - 29320. [Abstract] [Full Text] [PDF]

				A. Shainskaya and S. J. D. Karlish Chymotryptic Digestion of the Cytoplasmic Domain of the beta Subunit of Na/K-ATPase Alters Kinetics of Occlusion of Rb[IMAGE] Ions J. Biol. Chem., April 26, 1996; 271(17): 10309 - 10316. [Abstract] [Full Text] [PDF]

				P. A. Pedersen, J. H. Rasmussen, and P. L. Jørgensen Expression in High Yield of Pig alpha1beta1 Na,K-ATPase and Inactive Mutants D369N and D807N in Saccharomyces cerevisiae J. Biol. Chem., February 2, 1996; 271(5): 2514 - 2522. [Abstract] [Full Text] [PDF]

				R. A. Farley, S. Schreiber, S.-G. Wang, and G. Scheiner-Bobis A Hybrid between Na+,K+-ATPase and H+,K+-ATPase Is Sensitive to Palytoxin, Ouabain, and SCH 28080 J. Biol. Chem., January 19, 2001; 276(4): 2608 - 2615. [Abstract] [Full Text] [PDF]