The gamma subunit of Na+, K+-ATPase: role on ATPase activity and regulatory phosphorylation by PKA.
In kidney, Na+, K+-ATPase is an oligomer (alphabeta gamma) with equimolar amounts of essential alpha and beta subunits and one small hydrophobic FXYD protein (gamma subunit). This report describes gamma subunit as an activator of pig kidney outer medulla Na+, K+-ATPase in aqueous medium. The effects of gamma subunit on Na+, K+-ATPase were dose-dependent and preincubation-dependent. Changes in alphabeta/gamma stoichiometry did not alter Km1 for ATP, and slightly increased Km2, but Vmax was increased at both catalytic and regulatory sites. Hydroxylamine treatment of enzyme phosphorylated by ATP (E-P), in the presence of additional gamma subunit, revealed that 52% of the E-P accumulation was not via acyl-phosphate formation. The gamma subunit was phosphorylated by endogenous kinases and by commercial catalytic subunit of protein kinase A (PKA). Additionally, we demonstrated that PKA phosphorylation of gamma subunit increased its capacity to stimulate ATP hydrolysis. These results suggest that gamma subunit can act as an intrinsic Na+, K+-ATPase regulator in kidney.
Cortes
Vanessa Faria
VF
Instituto de Bioquímica Médica, Programa de Biologia Estrutural, Lab. LERPA-Estrutura e Regulação de Proteínas e ATPases, Alameda Bahuinia 400, prédio do CCS, bloco H2-026, Universidade Federal do Rio de Janeiro, CEP 21941-590, Rio de Janeiro, RJ, Brazil.
Veiga-Lopes
Fabio Eduardo
FE
Barrabin
Hector
H
Alves-Ferreira
Marcelo
M
Fontes
Carlos Frederico Leite
CF
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D007668:Kidney / Q000201:enzymology / Q000378:metabolism; D007700:Kinetics; D008055:Lipids / Q000737:chemistry / Q000302:isolation & purification / Q000494:pharmacology; D010042:Ouabain / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D020836:Protein Structure, Quaternary; D021122:Protein Subunits / Q000737:chemistry / Q000378:metabolism; D000254:Sodium-Potassium-Exchanging ATPase / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013552:Swine
D004791:Enzyme Inhibitors; D008055:Lipids; D021122:Protein Subunits; D010042:Ouabain; D000255:Adenosine Triphosphate; D017868:Cyclic AMP-Dependent Protein Kinases; D000254:Sodium-Potassium-Exchanging ATPase