Synergistic stimulation by potassium and ammonium of K+-phosphatase activity in gill microsomes from the crab Callinectes ornatus acclimated to low salinity: Novel property of a primordial pump

Abstract

We provide an extensive characterization of the modulation by p-nitrophenylphosphate, Mg2+, Na+, K+, Rb+, NH4+ and pH of gill microsomal K+-phosphatase activity in the posterior gills of Callinectes ornatus acclimated to low salinity (21‰). The synergistic stimulation by K+ and NH4+ of the K+-phosphatase activity is a novel finding, and may constitute a species-specific feature of K+/NH4+ interplay that regulates crustacean gill (Na+, K+)-ATPase activity. p-Nitrophenylphosphate was hydrolyzed at a maximum rate (V) of 69.2±2.8nmolPimin−1mg−1 with K0.5=2.3±0.1mmolL−1, obeying cooperative kinetics (nH=1.7). Stimulation by Mg2+ (V=70.1±3.0nmolPimin−1mg−1, K0.5=0.88±0.04mmolL−1), K+ (V=69.6±2.7nmolPimin−1mg−1, K0.5=1.60±0.07mmolL−1) and NH4+ (V=90.8±4.0nmolPimin−1mg−1, K0.5=9.2±0.3mmol L−1) all displayed site-site interaction kinetics. In the presence of NH4+, enzyme affinity for K+ unexpectedly increased by 7-fold, while affinity for NH4+ was 28-fold greater in the presence than absence of K+. Ouabain partially inhibited K+-phosphatase activity (KI=320±14.0μmolL−1), more effectively when NH4+ was present (KI=240±12.0μmolL−1). We propose a model for the synergistic stimulation by K+ and NH4+ of the K+-phosphatase activity of the (Na+, K+)-ATPase from C. ornatus posterior gill tissue.