RESUMEN
Polymyxin B, a cyclic peptide antibiotic, inhibits Ca2+-ATPase, p-nitrophenyl phosphatase and phosphorylase kinase activities associated with rabbit skeletal muscle sarcoplasmic reticulum membranes; 50% inhibition is induced by 100 microM, 130 microM and 550 microM of polymyxin respectively. The fluorescence intensity of fluorescein isothiocyanate-labeled Ca2+-ATPase, decreases in the presence of polymyxin (50% of the total decrease at 70 microM polymyxin). On the other hand, the polypeptide inhibits calmodulin-dependent endogenous phosphorylation of 60 kDa, 20 kDa and 14 kDa membrane proteins, while an increase of calmodulin-dependent phosphorylation is observed in 132 kDa and 86 kDa proteins.
Asunto(s)
Polimixina B/farmacología , Polimixinas/farmacología , Retículo Sarcoplasmático/efectos de los fármacos , 4-Nitrofenilfosfatasa/antagonistas & inhibidores , Animales , ATPasas Transportadoras de Calcio/antagonistas & inhibidores , Calmodulina/farmacología , Técnicas In Vitro , Membranas Intracelulares/efectos de los fármacos , Membranas Intracelulares/metabolismo , Proteínas de la Membrana/metabolismo , Fosforilasa Quinasa/antagonistas & inhibidores , Conejos , Retículo Sarcoplasmático/metabolismoRESUMEN
Nonactivated rabbit skeletal muscle phosphorylase kinase is inhibited by the polymyxins A, B, D and E when assayed at pH 8.6. Polymyxin B is the most effective inhibitor, causing 50% inhibition at 0.3 mM. Following the effect of polymyxin B on the kinase activity toward troponin, no inhibition was observed. In contrast, polymyxin B was found to greatly stimulate the autophosphorylation of phosphorylase kinase. About 10 mol of phosphate per tetramer (alpha beta nu delta) were incorporated in presence of polymyxin B (full autophosphorylation). This incorporation was about 6-fold higher than that observed without polymyxin. The stimulation of autophosphorylation by polymyxin B was accompanied with enhancement of the rate of autoactivation at pH 6.8.
Asunto(s)
Fosforilasa Quinasa/antagonistas & inhibidores , Polimixina B/farmacología , Polimixinas/farmacología , Animales , Activación Enzimática/efectos de los fármacos , Concentración de Iones de Hidrógeno , Músculos/enzimología , Fosforilasa Quinasa/metabolismo , Fosforilación , ConejosRESUMEN
A number of aliphatic amines was shown to stimulate AMP-dependent activity of phosphorylase b. The extent of stimulation depends on the molecular structure of amines. For linear amines, the longer the linear chain, the greater the stimulation observed. High concentrations of amines were able to induce a small activation of phosphorylase b in the absence of AMP. Kinetic studies of phosphorylase b indicated that the presence of n-hexylamine (a) results in lowering Km values for AMP and glucose 1-phosphate, (b) increases maximal velocity of the enzyme, and (c) modifies the glucose 6-phosphate, ATP, caffeine, and glucose binding sites of the enzyme by increasing the inhibition constants for these inhibitors. In contrast, the activity of phosphorylase b' is not altered by n-hexylamine. This fact suggests the possibility that amines interact with the N-terminal tail of phosphorylase b chain.