title: Gill (Na+,K+)-ATPase from the blue crab Callinectes danae: modulation of K+-phosphatase activity by potassium and ammonium ions.
abstract: The kinetic properties of a microsomal gill (Na(+),K(+))-ATPase from the blue crab Callinectes danae were analyzed using the substrate p-nitrophenylphosphate. The (Na(+),K(+))-ATPase hydrolyzed PNPP obeying cooperative kinetics (n=1.5) at a rate of V=125.4+/-7.5 U mg(-1) with K(0.5)=1.2+/-0.1 mmol l(-1); stimulation by potassium (V=121.0+/-6.1 U mg(-1); K(0.5)=2.1+/-0.1 mmol l(-1)) and magnesium ions (V=125.3+/-6.3 U mg(-1); K(0.5)=1.0+/-0.1 mmol l(-1)) was cooperative. Ammonium ions also stimulated the enzyme through site-site interactions (n(H)=2.7) to a rate of V=126.1+/-4.8 U mg(-1) with K(0.5)=13.7+/-0.5 mmol l(-1). However, K(+)-phosphatase activity was not stimulated further by K(+) plus NH(4)(+) ions. Sodium ions (K(I)=36.7+/-1.7 mmol l(-1)), ouabain (K(I)=830.3+/-42.5 micromol l(-1)) and orthovanadate (K(I)=34.0+/-1.4 nmol l(-1)) completely inhibited K(+)-phosphatase activity. The competitive inhibition by ATP (K(I)=57.2+/-2.6 micromol l(-1)) of PNPPase activity suggests that both substrates are hydrolyzed at the same site on the enzyme. These data reveal that the K(+)-phosphatase activity corresponds strictly to a (Na(+),K(+))-ATPase in C. danae gill tissue. This is the first known kinetic characterization of K(+)-phosphatase activity in the portunid crab C. danae and should provide a useful tool for comparative studies.
authors.affiliation: Departamento de Qui;mica, Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto, Universidade de São Paulo, 14040-901, SP, Ribeirão Preto, Brazil.
authors.forename: D C
authors.initials: DC
authors.lastname: Masui
authors.forename: R P M
authors.initials: RP
authors.lastname: Furriel
authors.forename: F L M
authors.initials: FL
authors.lastname: Mantelatto
authors.forename: J C
authors.initials: JC
authors.lastname: McNamara
authors.forename: F A
authors.initials: FA
authors.lastname: Leone
mesh_terms: D000818:Animals; D001665:Binding Sites; D003386:Brachyura / Q000201:enzymology*; D033364:Decapoda / Q000201:enzymology; D005880:Gills / Q000201:enzymology*; D007700:Kinetics; D008861:Microsomes / Q000201:enzymology; D009596:Nitrophenols / Q000378:metabolism; D009943:Organophosphorus Compounds / Q000378:metabolism; D010042:Ouabain / Q000494:pharmacology; D011188:Potassium / Q000494:pharmacology*; D000644:Quaternary Ammonium Compounds / Q000494:pharmacology*; D000254:Sodium-Potassium-Exchanging ATPase / Q000187:drug effects / Q000378:metabolism*; D014638:Vanadates / Q000494:pharmacology
chemical_list: D009596:Nitrophenols; D009943:Organophosphorus Compounds; D000644:Quaternary Ammonium Compounds; C008644:nitrophenylphosphate; D014638:Vanadates; D010042:Ouabain; D000254:Sodium-Potassium-Exchanging ATPase; D011188:Potassium