title: Identification of a crab gill FXYD2 protein and regulation of crab microsomal Na,K-ATPase activity by mammalian FXYD2 peptide.
abstract: This investigation discloses the recognition of an FXYD2 protein in a microsomal Na,K-ATPase preparation from the posterior gills of the blue crab, Callinectes danae, by a mammalian (rabbit) FXYD2 peptide specific antibody (γC(33)) and MALDI-TOF-TOF mass spectrometry techniques. This is the first demonstration of an invertebrate FXYD2 protein. The addition of exogenous pig FXYD2 peptide to the crab gill microsomal fraction stimulated Na,K-ATPase activity in a dose-dependent manner. Exogenous pig FXYD2 also considerably increased enzyme affinity for K(+), ATP and NH(4)(+). K(0.5) for Na(+) was unaffected. Exogenous pig FXYD2 increased the V(max) for stimulation of gill Na,K-ATPase activity by Na(+), K(+) and ATP, by 30% to 40%. The crab gill FXYD2 is phosphorylated by PKA, suggesting a regulatory function similar to that known for the mammalian enzyme. The PKA-phosphorylated pig FXYD2 peptide stimulated the crab gill Na,K-ATPase activity by 80%, about 2-fold greater than did the non-phosphorylated peptide. Stimulation by the PKC-phosphorylated pig FXYD2 peptide was minimal. These findings confirm the presence of an FXYD2 peptide in the crab gill Na,K-ATPase and demonstrate that this peptide plays an important role in regulating enzyme activity.
authors.affiliation: Instituto de Bioquímica Médica, Laboratório de Estrutura e Regulação de Proteínas e ATPases, Programa de Biologia Estrutural, RJ, Brazil.
authors.forename: Elias C C
authors.initials: EC
authors.lastname: Silva
authors.forename: Douglas C
authors.initials: DC
authors.lastname: Masui
authors.forename: Rosa P
authors.initials: RP
authors.lastname: Furriel
authors.forename: John C
authors.initials: JC
authors.lastname: McNamara
authors.forename: Hector
authors.initials: H
authors.lastname: Barrabin
authors.forename: Helena M
authors.initials: HM
authors.lastname: Scofano
authors.forename: Jonas
authors.initials: J
authors.lastname: Perales
authors.forename: André
authors.initials: A
authors.lastname: Teixeira-Ferreira
authors.forename: Francisco A
authors.initials: FA
authors.lastname: Leone
authors.forename: Carlos Frederico L
authors.initials: CF
authors.lastname: Fontes
mesh_terms: D000818:Animals; D003445:Crustacea / Q000378:metabolism*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D005880:Gills / Q000201:enzymology*; D006801:Humans; D008861:Microsomes / Q000201:enzymology*; D010766:Phosphorylation; D000254:Sodium-Potassium-Exchanging ATPase / Q000378:metabolism*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013552:Swine / Q000378:metabolism
chemical_list: D017868:Cyclic AMP-Dependent Protein Kinases; C416502:FXYD2 protein, human; D000254:Sodium-Potassium-Exchanging ATPase