Characterization of intestinal Na+-K+-ATPase in the gilthead seabream (Sparus aurata L.). Evidences for tissue-specific heterogeneity.

Abstract

Gilthead seabream intestine contains both a Mg2+-dependent Na+–K+-ATPase which is completely inhibited by 1×10−3 M ouabain, and also a residue-ATPase activity that is entirely ouabain-insensitive. The maximal activity of intestinal Na+–K+-ATPase (35.15 μmol inorganic phosphate (Pi) mg protein−1 h−1) was observed in the microsomal fraction at 35°C, pH 7.5, 2–5 mM MgCl2, 5 mM ATP, 10 mM K+ and 200 mM Na+. The intestinal Na+–K+-ATPase of gilthead seabream exhibits similar characteristics to other teleost Na+–K+-ATPases regarding pH dependence, Mg2+/ATP optimal ratios and ouabain sensitivity, but exhibits unusual sensitivity to ionic strength and cation promoted cooperative activation and higher affinity for magnesium and ATP when compared to other marine and euryhaline teleosts. The Arrhenius plot for intestinal Na+–K+-ATPase showed a break point at 15.41°C, with similar activation energies above and below the discontinuity point. The analysis of polar lipid fatty acid composition was correlated to the break point in the Arrhenius plot, suggesting a regulatory role of the lipid microenvironment on the enzyme activity. Finally, the kinetic characteristics, temperature-activity relationship, fatty acid composition and substrate dependence of intestinal Na+–K+-ATPase are compared with literature data and discussed on the basis of the differences between species and also between osmoregulatory tissues. It is concluded that the Na+–K+-ATPase of Sparus aurata is differently and specifically expressed between the osmoregulatory organs.