Leone Francisco A, Bezerra Thais M S, Garçon Daniela P, Lucena Malson N, Pinto Marcelo R, Fontes Carlos F L, McNamara John C
Departamento de Química, Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP, Brasil.
Departamento de Biologia Molecular, Centro de Ciências Exatas e da Natureza, Universidade Federal da Paraíba, João Pessoa, PB, Brasil.
PLoS One. 2014 Feb 21;9(2):e89625. doi: 10.1371/journal.pone.0089625. eCollection 2014.
We investigate the synergistic stimulation by K(+) plus NH4 (+) of (Na(+), K(+))-ATPase activity in microsomal preparations of whole zoea I and decapodid III, and in juvenile and adult river shrimp gills. Modulation of (Na(+), K(+))-ATPase activity is ontogenetic stage-specific, and particularly distinct between juveniles and adults. Although both gill enzymes exhibit two different sites for K(+) and NH4 (+) binding, in the juvenile enzyme, these two sites are equivalent: binding by both ions results in slightly stimulated activity compared to that of a single ionic species. In the adult enzyme, the sites are not equivalent: when one ion occupies its specific binding site, (Na(+), K(+))-ATPase activity is stimulated synergistically by ≈ 50% on binding of the complementary ion. Immunolocalization reveals the enzyme to be distributed predominantly throughout the intralamellar septum in the gill lamellae of juveniles and adults. Western blot analyses demonstrate a single immunoreactive band, suggesting a single (Na(+), K(+))-ATPase α-subunit isoform that is distributed into different density membrane fractions, independently of ontogenetic stage. We propose a model for the modulation by K(+) and NH4 (+) of gill (Na(+), K(+))-ATPase activity. These findings suggest that the gill enzyme may be regulated by NH4 (+) during ontogenetic development in M. amazonicum.
我们研究了钾离子(K⁺)与铵离子(NH₄⁺)对整个第一期蚤状幼体和第三期大眼幼体的微粒体制备物,以及幼体和成体河虾鳃中(钠钾)-ATP酶活性的协同刺激作用。(钠钾)-ATP酶活性的调节具有个体发育阶段特异性,在幼体和成体之间尤为明显。虽然两种鳃酶都表现出钾离子和铵离子的两个不同结合位点,但在幼体酶中,这两个位点是等效的:与单一离子种类相比,两种离子结合都会导致活性略有刺激。在成体酶中,这些位点不等效:当一种离子占据其特异性结合位点时,(钠钾)-ATP酶活性在互补离子结合时会协同刺激约50%。免疫定位显示该酶主要分布在幼体和成体鳃小片的板内间隔中。蛋白质免疫印迹分析显示有一条免疫反应带,表明存在单一的(钠钾)-ATP酶α亚基同工型,其分布在不同密度的膜组分中,与个体发育阶段无关。我们提出了一个钾离子和铵离子对鳃(钠钾)-ATP酶活性进行调节的模型。这些发现表明,在亚马逊沼虾的个体发育过程中,鳃酶可能受铵离子调节。