RESUMO
Recent studies have provided evidence for a role of cyclic ADP-ribose (cADPR) in the regulation of intracellular calcium in smooth muscles of the intestine, blood vessels and airways. We investigated the presence and subcellular localization of ADP-ribosyl cyclase, the enzyme that catalyzes the conversion of beta-NAD(+) to cADPR, and cADPR hydrolase, the enzyme that degrades cADPR to ADPR, in tracheal smooth muscle (TSM). Sucrose density fractionation of TSM crude membranes provided evidence that ADP-ribosyl cyclase and cADPR hydrolase activities were associated with a fraction enriched in 5'-nucleotidase activity, a plasma membrane marker enzyme, but not in a fraction enriched in either sarcoplasmic endoplasmic reticulum calcium ATPase or ryanodine receptor channels, both sarcoplasmic reticulum markers. The ADP-ribosyl cyclase and cADPR hydrolase activities comigrated at a molecular weight of approximately 40 kDa on SDS-PAGE. This comigration was confirmed by gel filtration chromatography. Investigation of kinetics yielded K(m) values of 30.4+/-1.5 and 695. 3+/-171.2 microM and V(max) values of 330.4+/-90 and 102.8+/-17.1 nmol/mg/h for ADP-ribosyl cyclase and cADPR hydrolase, respectively. These results suggest a possible role for cADPR as an endogenous modulator of [Ca(2+)](i) in porcine TSM cells.
Assuntos
Carbono-Oxigênio Liases/metabolismo , Músculo Liso/enzimologia , Fósforo-Oxigênio Liases/metabolismo , Traqueia/enzimologia , ADP-Ribosil Ciclase , Animais , Western Blotting , Fracionamento Celular , Cromatografia em Gel , Eletroforese em Gel de Poliacrilamida , Cinética , Músculo Liso/ultraestrutura , Radioisótopos de Fósforo , Espectrometria de Fluorescência , Suínos , Traqueia/ultraestruturaAssuntos
Aplysia/enzimologia , Fluorometria/métodos , Nucleotídeos de Guanina/metabolismo , Liases/análise , N-Glicosil Hidrolases/análise , NAD/análogos & derivados , ADP-Ribosil Ciclase , Animais , Feminino , Hidrólise , Nucleotídeos de Inosina/metabolismo , Liases/isolamento & purificação , Masculino , Complexos Multienzimáticos/análise , Complexos Multienzimáticos/isolamento & purificação , N-Glicosil Hidrolases/isolamento & purificação , NAD/metabolismo , Ovário/enzimologia , Testículo/enzimologiaAssuntos
Antígenos CD , Antígenos de Diferenciação/biossíntese , N-Glicosil Hidrolases/biossíntese , ADP-Ribosil Ciclase , ADP-Ribosil Ciclase 1 , Antígenos de Diferenciação/genética , Clonagem Molecular , Fermentação , Vetores Genéticos , Glicerol/metabolismo , Humanos , Glicoproteínas de Membrana , N-Glicosil Hidrolases/genética , Pichia/genética , Pichia/crescimento & desenvolvimento , Proteínas Recombinantes/biossíntese , SolubilidadeRESUMO
A comparison of two methods of selenium application, banding and foliar spray, of sodium selenite (Na2SeO3) on total glycoalkaloid (TGA) and nitrate nitrogen (NO3-N) was studied during each of two consecutive years. The levels of application used were 0.0, 1.6 (0.75), 3.36 (1.5), and 5.6 (2.5) kg/ha (ppm soil). Both TGA and NO3-N were significantly reduced by application of 1.5 and 2.5 ppm of sodium selenite. Tuber selenium levels were significantly increased at all levels of application, using either banding or foliar spray, but were well below the toxic range for human consumption. Banding resulted in greater uptake of Se, and greater decreases in TGA and NO3-N as compared to foliar spray.
