Structural organization of alpha-subunit from purified and microsomal toad kidney (Na+ + K+)-ATPase as assessed by controlled trypsinolysis

Détails

ID Serval
serval:BIB_BBEA2D1C96EB
Type
Article: article d'un périodique ou d'un magazine.
Collection
Publications
Institution
Titre
Structural organization of alpha-subunit from purified and microsomal toad kidney (Na+ + K+)-ATPase as assessed by controlled trypsinolysis
Périodique
Biochimica et Biophysica Acta (BBA) - Biomembranes
Auteur⸱e⸱s
Zamofing Danièle, Rossier Bernard C., Geering Käthi
ISSN
0005-2736
ISSN-L
1879-2642
Statut éditorial
Publié
Date de publication
11/1987
Peer-reviewed
Oui
Volume
904
Numéro
2
Pages
381-391
Langue
anglais
Notes
Publication types: Journal Article ; Research Support, Non-U.S. Gov't - Publication Status: ppublish
Résumé
The membrane organization of the alpha-subunit of purified (Na+ + K+)-ATPase ((Na+ + K+)-dependent adenosine triphosphate phosphorylase, EC 3.6.1.3) and of the microsomal enzyme of the kidney of the toad Bufo marinus was compared by using controlled trypsinolysis. With both enzyme preparations, digestions performed in the presence of Na+ yielded a 73 kDa fragment and in the presence of K+ a 56 kDa, a 40 kDa and small amounts of a 83 kDa fragment from the 96 kDa alpha-subunit. In contrast to mammalian preparations (Jørgensen, P.L. (1975) Biochim. Biophys. Acta 401, 399-415), trypsinolysis of the purified amphibian enzyme led to a biphasic loss of (Na+ + K+)-ATPase activity in the presence of both Na+ and K+. These data could be correlated with an early rapid cleavage of 3 kDa from the alpha-subunit in both ionic conditions and a slower degradation of the remaining 93 kDa polypeptide. On the other hand, in the microsomal enzyme, a 3 kDa shift of the alpha-subunit could only be produced in the presence of Na+. Our data indicate that (1) purification of the amphibian enzyme with detergent does not influence the overall topology of the alpha-subunit but produces a distinct structural alteration of its N-terminus and (2) the amphibian kidney enzyme responds to cations with similar conformational transitions as the mammalian kidney enzyme. In addition, anti alpha-serum used on digested enzyme samples revealed on immunoblots that the 40 kDa fragment was better recognized than the 56 kDa fragment. It is concluded that the NH2-terminal of the alpha-subunit contains more antigenic sites than the COOH-terminal domain in agreement with the results of Farley et al. (Farley, R.A., Ochoa, G.T. and Kudrow, A. (1986) Am. J. Physiol. 250, C896-C906).
Mots-clé
Animals, Bufo marinus, Intracellular Membranes, Kidney, Kinetics, Macromolecular Substances, Microsomes, Molecular Weight, Peptide Fragments, Protein Conformation, Sodium-Potassium-Exchanging ATPase, Trypsin, Biophysics, Cell Biology, Biochemistry
Pubmed
Web of science
Création de la notice
24/01/2008 13:00
Dernière modification de la notice
20/08/2019 15:29
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