Assuntos
Bordetella pertussis/fisiologia , AMP Cíclico/metabolismo , Inibidores de Adenilil Ciclases , Adenilil Ciclases/isolamento & purificação , Adenilil Ciclases/metabolismo , Glândulas Suprarrenais/efeitos dos fármacos , Animais , Proteínas de Bactérias/isolamento & purificação , Proteínas de Bactérias/metabolismo , Toxinas Bacterianas/farmacologia , Bordetella pertussis/enzimologia , Cálcio/farmacologia , Calmodulina/farmacologia , Bovinos , Endotoxinas/farmacologia , Ativação Enzimática/efeitos dos fármacos , Humanos , Ratos , Esteroides/biossíntese , Fatores de Virulência de BordetellaRESUMO
A variety of proteins and tissue preparations (rabbit erythrocyte lysate, catalase, peroxidase, creatine phosphokinase, and lima bean trypsin inhibitor) contain protein activator(s) of the extracellular adenylate cyclase of intact Bordetella pertussis organisms. Stimulation of adenylate cyclase activity of up to 1000-fold over basal activity can be obtained. Activation of the adenylate cyclase is due to the presence of calmodulin in these protein preparations. The criteria to establish this were: Ca2+ dependence of the activation, inhibition by trifluoperazine, heat stability of the activator, chromatographic behavior like authentic calmodulin, and stimulation of cyclic nucleotide phosphodiesterase by the activators. The great sensitivity of the B.pertussis adenylate cyclase assay makes this and ideal system for the detection of trace amounts of calmodulin, in the presence of large amounts of other proteins.
Assuntos
Adenilil Ciclases/metabolismo , Bordetella pertussis/enzimologia , Proteínas/farmacologia , Animais , Proteínas Sanguíneas/farmacologia , Catalase/farmacologia , Ativação Enzimática , Cinética , Proteínas/análise , Coelhos , Inibidores da Tripsina/farmacologiaRESUMO
The adenylate cyclase of Bordetella pertussis is stimulated 100- to 1000-fold in a dose-dependent manner by calf brain calmodulin. The system has the following properties. (i) The activation is prevented by ethylene glycol bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid and restored by Ca2+. (ii) Oxidation of the methionine residues of calmodulin abolishes the ability to activate the cyclase. (iii) Trifluoperazine inhibits calmodulin-activated cyclase. (iv) A troponin C preparation stimulates the B. pertussis cyclase with < 0.01 the potency of calmodulin. Although calmodulin has not been demonstrated in prokaryotes, this is an example of a (eukaryotic) calmodulin effect in a prokaryote.
Assuntos
Adenilil Ciclases/metabolismo , Bordetella pertussis/enzimologia , Proteínas de Ligação ao Cálcio/farmacologia , Calmodulina/farmacologia , Cálcio/metabolismo , Calmodulina/antagonistas & inibidores , Ácido Egtázico/farmacologia , Ativação Enzimática/efeitos dos fármacos , Troponina/farmacologiaRESUMO
Apparent values of Km and Vmax have been measured for catalysis of hydrolysis of unsonicated egg lecithin liposomes, activated through addition of 0.4 M n-hexanol, by phospholipases A2 from bee and snake venoms and by phospholipase C from Clostridium welchii as a function of the concentration of three surfactants: hexadecylamine, hexadecyltrimethylammonium bromide, and dihexadecyl phosphate. For all three enzymes, values of Km and Vmax show little or no dependence on the concentration of these ionic surfactants, demonstrating that the liposomal surface charge is not a crucial factor in determining susceptibility to phospholipase-catalyzed hydrolysis.
