SEARCH · Search PubMed

Results for “PHOSPHORYLASES”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

SourceResearch connectionFrom yearThrough year

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 829 records · Page 46Linked to original sources

PubMed1982

5-benzylacyclouridine and 5-benzyloxybenzylacyclouridine, potent inhibitors of uridine phosphorylase.

Various pyrimidine acyclonucleosides (1-(2'-hydroxyethoxymethyl)uracils) are specific inhibitors of uridine phosphorylase[Niedzwicki et al., Biochem. Pharmac. 30, 2097 (1981) )). 5-Benzyluracils have also been shown to inhibit this enzyme[Baker and Kelley, J. med. Chem. 13, 461 (1970); Woodman et al., Biochem. Pharmac. 29, 1059 (1980) )). We have synthesized the acyclonucleoside analogs of 5-benzyluracil (BU) and 5-benzyloxybenzyluracil (BBU). These compounds, 5-benzyl-1-(2'-hydroxyethoxymethyl)uracil (BAU) and 5-(m-benzyloxybenzyl)-1-(2'-hydroxyethoxymethyl)uracil (BBAU), are potent inhibitors of uridine phosphorylase. K1 values of 98 and 32 nM were estimated for BAU and BBAU respectively. These compounds are better inhibitors of uridine phosphorylase than BU (K1= 1575 nM), BBU (K1=270 nM), and all other compounds previously tested, and they have no effect on thymidine phosphorylase, uridine-cytidine kinase, or thymidine kinase. Potential chemotherapeutic applications of BAU and BBAU are discussed.

Animals↗

PubMed1985

Salvage of uridine in the mouse. Effect of uridine phosphorylase pretreatment.

Salvage of circulating nucleosides provides an alternative to de novo synthesis of nucleotides and may modify response to antimetabolites. We have investigated treatment with uridine phosphorylase as a means of inhibiting salvage of uridine in vivo. Examination of the metabolism of intravenous [3H] uridine in mice revealed that 30-40% was salvaged by conversion to uracil nucleotides and the remainder was catabolized. In contrast, less than 0.3% of intravenous [3H]uracil was salvaged. Addition of partially purified bacterial uridine phosphorylase to plasma produced a rapid phosphorolysis of uridine. In vivo, 1.5 hr after intravenous injection of 9 units of uridine phosphorylase, plasma activity (1.3 units/ml) was 65-fold greater than that of control mice. Pretreatment with uridine phosphorylase prior to administration of [3H]uridine produced a marked (65-92%) but incomplete inhibition of salvage of uridine in all tissues examined. The dose required to produce 50% inhibition of uridine salvage at 1 hr was 2 to 2.5 units/mouse. The inhibition of nucleoside salvage by this approach may permit an evaluation of the role of nucleoside salvage in the supply of cellular nucleotides and the effects of concurrent inhibition of de novo and salvage pathways for nucleotide synthesis.

Animals↗

PubMed1983

Adenosine phosphorylase activity in mycoplasma-free growth media for mammalian cells.

Mammalian cells have enzymes that deaminate adenosine to inosine, which can readily be phosphorolysed to hypoxanthine. They do not, however, possess enzymes to form adenine by the cleavage of adenosine. For this reason, the release of adenine from adenosine by mammalian cell cultures has usually been interpreted as indicating the presence of mycoplasma, a frequent microbial contaminant that contains high levels of adenosine phosphorylase. We found that some human lymphoblast cultures free of mycoplasma showed high levels of adenosine cleavage and that this activity resulted from adenosine phosphorylase in the bovine serum used as the culture growth supplement. A survey of 13 serum supplements disclosed that fetal bovine serum (six lots) contains the highest adenosine phosphorylase activity, ranging from 9 to 648 nmol adenine produced per hour per ml serum; newborn calf serum (four lots) has much less activity, ranging from 0 to 5 nmol adenine produced per hour per ml serum; and donor horse serum (three lots) contains no detectable activity. These results suggest that mycoplasma tests dependent on the presence of adenosine phosphorylase or other enzyme activities may give false-positives with cultures containing fetal bovine serum supplements.

Adenosine↗

PubMed1984

The Mg2+ requirements of nonactivated and activated rat liver phosphorylase kinase. Inhibition of the activated form by free Mg2+.

Incubation of rat liver phosphorylase kinase in the presence of MgATP results in a time-dependent increase in activity, i.e., activation. Determination of the magnitude of activation depends, in large part, on the relative concentrations of Mg2+ and ATP used in the phosphorylase kinase activity assay, such that as the Mg2+ to ATP ratio increases less activation is detectable. Prior to activation, maximal activity of nonactivated phosphorylase kinase requires a 2-3-fold molar excess of Mg2+ (i.e., free Mg2+) over ATP. MgATP-dependent activation of the enzyme results in an alteration in the free Mg2+ requirement such that the activity of the activated enzyme is sharply inhibited by the free cation. Inhibition by free Mg2+ of the activated enzyme is rapidly reversed by removal of free Mg2+ but is not affected by addition of Ca2+. Both nonactivated and activated forms of the enzyme appear to be inhibited by free ATP4-. The results show that the use of high concentrations of free Mg2+ in the phosphorylase kinase activity assay can blunt or completely obscure changes in enzyme activity following activation of the enzyme.

Adenosine Triphosphate↗

PubMed1986

The diurnal rhythm of liver glycogen phosphorylase: correlating changes in enzyme activity and enzymic protein.

The diurnal rhythm of phosphorylase activity in mouse liver extracts was correlated with the 24 h fluctuations in phosphorylase protein. This last measurement was made using rocket immunoelectrophoresis. The peak activity of phosphorylase appeared coincident (at 18:00 h) with the greatest amount of phosphorylase protein detected. Conversely, the lowest activity measured and lowest enzymic protein content both occurred at 02:00 h. Regression analysis revealed a significant positive correlation between enzyme activity and protein. Thus changes in the cellular concentration of this enzyme are implicated in the diurnal rhythm of liver glycogen.

Animals↗

PubMed1987

Antigen reversion of glycogen phosphorylase isoenzyme in carcinoma and proliferative zone of intestinal metaplasia of the human stomach. An immunohistochemical study.

Specific antibodies to three purified glycogen phosphorylase isoenzymes, i.e., brain (or fetal), muscle, and liver types, were used to study the localization of these isoenzymes in carcinoma and the proliferative zone of intestinal metaplasia of the human stomach. Both the malignant cells of well-differentiated adenocarcinoma and the proliferative zone of some intestinal metaplasia of the stomach were stained when the antibrain-type phosphorylase isoenzyme antibody was used, but not when the other two types were used. The results suggest that brain-type phosphorylase in gastric carcinoma could be one example of fetal protein expression in cancer, and that the proliferative zone of some intestinal metaplasia having brain-type phosphorylase may histogenetically relate to well-differentiated adenocarcinoma.

Adenocarcinoma↗

PubMed1984

Ca2+-dependent stimulation of muscle and liver phosphorylase kinase by heparin.

Heparin stimulates the activity of nonactivated and activated skeletal muscle phosphorylase kinase in a Ca2+-dependent manner. The stimulatory effect of heparin on the activity of nonactivated phosphorylase kinase is also expressed in the presence of calmodulin and glycogen. Heparin acted in synergism with glycogen. Heparin increases the affinity of phosphorylase kinase to Ca2+ 5-12 fold depending upon the activation conditions. Ca2+ influences the stimulation of liver phosphorylase kinase by heparin in a similar way.

Animals↗

PubMed1989

Further evaluation of cofactor as a turnover label for glycogen phosphorylase.

1. The cofactor of glycogen phosphorylase, pyridoxal phosphate (PLP), is stably associated with the enzyme and has been used as a label in the determination of the turnover of the skeletal muscle enzyme in vivo. 2. Mice were injected with radiolabelled pyridoxine that was subsequently converted to PLP and incorporated into phosphorylase. 3. In this study we have resolved phosphorylase-bound label from that associated with the other PLP-containing enzymes and free label by affinity and size-exclusion chromatography. 4. The decay of radioactive pools was assessed after an extended period post-injection to minimize the effects of isotope reutilization. 5. These modifications have allowed refinement of our previous estimate of the rate of degradation of muscle phosphorylase.

Animals↗

PubMed1990

Role of calcium and cyclic AMP on the activation of lymphocyte glycogen phosphorylase by mitogens.

1. Various mitogens such as concanavalin A, phytohaemagglutinin, the pokeweed mitogen and trypsin were found to produce a rapid and transient activation of glycogen phosphorylase activity of lymphocytes incubated in a Krebs-Ringer-bicarbonate-glucose buffer. 2. Activation of the enzyme by these mitogens was always accompanied by an increase in the intracellular cyclic AMP concentration. 3. The presence of calcium ions in the incubation buffer was essential for obtaining the mitogen effects. Addition of ionophore A-23187 also produced an activation of glycogen phosphorylase, similar to that found in mitogen activation but without increase in intracellular cyclic AMP concentration. Dibutyril cyclic AMP also produced lymphocyte phosphorylase activation, even in the absence of extracellular calcium ions. 4. It is proposed that phosphorylase activation by mitogens occurs through a mechanism that involves the participation of both calcium ions and cyclic AMP.

Animals↗

PubMed1985

Two-dimensional electron microscopic analysis of the chalice form of phosphorylase kinase.

Phosphorylase kinase (Mr 1.3 X 10(6], a Ca2+-calmodulin-dependent protein kinase, plays a key role in the initiation of glycogenolysis. After purification on hydroxylapatite, the negatively stained enzyme was used for electron microscopy. In electron micrographs, phosphorylase kinase shows two major molecular forms: a butterfly form (approx. 60%) and a chalice form (approx. 40%). Images of the chalice form of the enzyme were computer-averaged by the method of single particle averaging. The following apparent molecular dimensions were obtained from the averages: total height, 20 nm; maximal width, 18 nm. The chalice form of phosphorylase kinase consists of a major structure termed the cup (11 nm X 18 nm), containing a large accessible cleft, and a minor structure termed the stem (8 nm X 9 nm). A closer examination of the images by averaging of molecular parts revealed two subpopulations of the cup part: a flexed (closed) type and an extended (open) type. The orifice, which can be closed partly by two protrusions (I, I'), is about 6 nm wide when the protrusions are flexed and 9 nm wide when they are extended. It is suggested that the substrates, e.g. phosphorylase b, may be accommodated in the large cleft of the enzyme. While the orientation of the protrusions (I, I') is the most obvious difference between the two types, more structural differences can be detected, suggesting a concerted movement of the protein domains against each other.

Animals↗

PubMed1986

Age-related change in adrenergic regulation of glycogen phosphorylase in rat hepatocytes.

The effects of development on adrenergic regulation of glycogenolysis were studied in rat liver. In isolated hepatocytes, the activation of glycogen phosphorylase by alpha-adrenergic stimulation decreased moderately with advancing age. However, activation by beta-adrenergic receptors more markedly declined and almost disappeared in isolated hepatocytes from 6-month-old rats. Furthermore, the ability to glucagon to stimulate phosphorylase activity and cAMP generation was found to decrease with increasing age. The age-related changes in the pattern of stimulation of glycogen phosphorylase by catecholamines in isolated hepatocytes were accompanied by parallels changes in the numbers of alpha 1- and beta-adrenoceptors in membranes prepared from the isolated hepatocytes. A progressive decrease in the total number of alpha 1-receptors measured with [3H]-prazosin and beta-receptors measured with [125I]cyanopindolol (CYP) was found with increasing age. The loss of beta-adrenergic receptors was much more dramatic. Our results suggest that age-related alterations of hepatic glycogen phosphorylase activation by catecholamines may in part be explained by the changes in the expression adrenergic receptors.

Aging↗

PubMed1988

Characterization of glycogen-synthase phosphatase and phosphorylase phosphatase in subcellular liver fractions.

Upon fractionation of a postmitochondrial supernatant from rat liver, the synthase phosphatase (EC 3.1.3.42) activity (assayed at high tissue concentrations) was largely recovered in the glycogen fraction and to a minor extent in the cytosol. In contrast, the phosphorylase phosphatase (EC 3.1.3.17) activity was approximately equally distributed between these two fractions, a lesser amount being recovered in the microsomal fraction. The phosphatase activities in the microsomal and glycogen fractions were almost completely inhibited by a preincubation with the modulator protein, a specific inhibitor of type-1 (ATP,Mg-dependent) protein phosphatases. In the cytosolic fraction, however, type-2A (polycation-stimulated) phosphatase(s) contributed significantly to the dephosphorylation of phosphorylase and of in vitro phosphorylated muscular synthase. Liver synthase b, used as substrate for the measurement of synthase phosphatase throughout this work, was only activated by modulator-sensitive phosphatases. Trypsin treatment of the subcellular fractions resulted in a dramatically increased (up to 1000-fold) sensitivity to modulator, a several-fold increase in phosphorylase phosphatase activity and a complete loss of synthase phosphatase activity. Similar changes occurred during dilution of the glycogen-bound enzyme. A preincubation with the deinhibitor protein, which is known to counteract the effects of inhibitor-1 and modulator, increased several-fold the phosphorylase phosphatase activity, but exclusively in the cytosolic and microsomal fractions. It did not affect the synthase phosphatase activity. Taken together, the results indicate the existence of distinct, multi-subunit type-1 phosphatases in the cytosolic, microsomal and glycogen fractions.

Animals↗

PubMed1993

Differential effects of calyculin A and okadaic acid on the glucose-induced regulation of glycogen synthase and phosphorylase activities in cultured hepatocytes.

The effects of the phosphatase inhibitors calyculin A and okadaic acid were investigated to determine the roles of protein phosphatases type 1 and 2A in the regulation of the activities of glycogen synthase and phosphorylase by glucose in a primary culture of hepatocytes. Glycogen synthesis, as measured by the incorporation of labelled glucose into glycogen, was inhibited in a dose-dependent manner by calyculin A (IC50 = 2.2 nM) and okadaic acid with (IC50 = 14 nM). Glucose-induced activation of glycogen synthase was inhibited by calyculin A and okadaic acid with IC50 values of 3.7 nM and 90 nM, respectively. Phosphorylase was simultaneously activated by these inhibitors with calyculin A again being more active (P < 0.001) than okadaic acid. The differing potencies (P < 0.001) of these inhibitors on the activities of glycogen synthase and phosphorylase were also observed with varying concentrations of glucose (5.6-60 mM) in the medium and at different incubation periods upto 120 min. It has been previously shown that both inhibitors inhibit protein phosphatase-2A with equal potency and calyculin A is a more potent inhibitor of protein phosphatase-1 than okadaic acid. Heat- and proteinase-treated cytosolic fractions from hepatocytes incubated with calyculin A and okadaic acid showed similar differential inhibitory activities towards purified types 1 and 2-A protein phosphatases. Hence, these data provide further evidence that protein phosphatase type-1 plays a major role in the control of glycogen synthesis by regulating the activities of glycogen synthase and phosphorylase.

Animals↗

PubMed1989

Human brain glycogen phosphorylase: characterization of fetal cDNA and genomic sequences.

Glycogen phosphorylase (alpha-1,4-glucan:orthophosphate D-glucosyltransferase, EC 2.4.1.1) is the rate-determining enzyme catalyzing glycogen degradation. Human brain has been demonstrated previously to express genes of both the liver and muscle isozymes of glycogen phosphorylase. In this report, a human fetal brain cDNA and genomic DNA corresponding to the brain isozyme of glycogen phosphorylase were isolated and characterized. Transcripts corresponding to this isozyme are present in human adult and fetal brain, and at low levels in other human fetal tissues. The predicted C-terminal sequence of the protein encoded by this cDNA and gene differ from that encoded by a phosphorylase cDNA isolated from a human astrocytoma cell line.

Amino Acid Sequence↗

PubMed1993

Effect of flavins on the rate of proteolytic digestion of muscle glycogen phosphorylase b.

The kinetics of tryptic proteolysis of rabbit skeletal muscle phosphorylase b has been registered by the diminishing of protein fluorescence intensity at lambda = 335 nm (excitation at 290 nm) or by the disappearance of the enzyme activity (0.02 M Hepes buffer, pH 6.8, 37 degrees C). The first procedure showed that flavins (riboflavin, FMN, FAD) protected the enzyme against tryptic digestion. Microscopic dissociation constants for the complexes of phosphorylase b with riboflavin, FMN and FAD were calculated from dependences of the initial digestion rate on the flavin concentration. They where equal to 30 +/- 1, 15.8 +/- 0.2 and 36 +/- 1 microM, respectively. No influence of FMN on the rate of the tryptic hydrolysis of phosphorylase b was observed when using the second procedure (enzyme activity test). FMN completely prevents the formation of 69-, 81- and 85-kDa fragments during 20 min incubation of phosphorylase b with trypsin.

Animals↗

PubMed1988

The Fourier transforms of the laser-induced absorption decay from glycogen phosphorylase and DOPA decarboxylase.

Fourier analysis of the laser-induced absorption decay curves of 3,4-dihydroxyphenylalanine (DOPA) decarboxylase and glycogen phosphorylase demonstrates a powerful technique in the analysis of complicated decay behavior. Phosphorylase which uses the pyridoxal 5'-phosphate cofactor in an unknown manner exhibits over weak absorption an intense decay while decarboxylase demonstrates only weak absorption. Fourier analysis of the decay curves clearly shows that phosphorylase has an intense absorption decay in the midst of three weaker ones and that decarboxylase only has three weak decays. This conclusion justifies the isolation and use of the intense decay of phosphorylase as an observable in the study of protein dynamics at the active site about the cofactor. The decay has demonstrated a movement of positive charge to substrate in the mechanism of phosphorylation of glycogen units.

Aromatic-L-Amino-Acid Decarboxylases↗

PubMed1989

Simulation of the activation of fat body glycogen phosphorylase and trehalose synthesis by peptide hormones in the American cockroach.

A model is described which simulates activation of glycogen phosphorylase and induction of trehalose synthesis in the fat body of the American cockroach, Periplaneta americana, by two hypertrehalosaemic peptides. Parameters for the model were estimated from literature data (Siegert et al., Insect Biochem. 16, 365), with the exception of the half-life of the physiologically active peptides, which was estimated from the model. The model describes satisfactorily the activation of glycogen phosphorylase and the increase of haemolymph carbohydrates, which is dependent on the activation of glycogen phosphorylase in the model. It further explains the observed differences in sensitivity for glycogen phosphorylase activation and increase in haemolymph carbohydrates by these peptides. Best fits were obtained with a physiological half-life of about 12-15 min for the peptides. This value is similar to what can be calculated from the in vivo effects of these peptides on heart beat (Gersch et al., Zool. Jb. Physiol. 86, 17), but it is considerably shorter than the published half-life of 1 h for radioactive peptide (Skinner et al., Insect Biochem. 17, 433). However, both values are compatible if part of the peptides in the haemolymph is not present in freely dissolved form, but bound to a haemolymph component. The model half-life would then represent the half-life of the free, physiologically active peptide, which was estimated from the disappearance of radioactive peptide to be about 12-15 min. This suggests, that the model half-life is realistic and physiologically more important than the half-life of radioactive peptide of 1 h.

Animals↗

PubMed1996

Calcium-dependent action of hypertrehalosemic hormone on activation of glycogen phosphorylase in cockroach fat body.

Signaling mechanisms for Blaberus discoidalis hypertrehalosemic hormone (Bld-HrTH)-dependent glycogen phosphorylase activation were investigated in vitro using fat body of the tropical cockroach, B. discoidalis. Brief treatment of fat bodies with Bld-HrTH in the absence of extracellular Ca2+ showed a significant activation of phosphorylase. Although extracellular Ca2+ was required for a full activation of phosphorylase by Bld-HrTH, stimulation in the absence of extracellular Ca2+ suggested that intracellular Ca2+ was also involved in Bld-HrTH signal transduction. Thapsigargin and thimerosal mobilize Ca2+ from intracellular stores by different mechanisms, and both chemicals stimulated phosphorylase activities as effectively as a maximum dose of Bld-HrTH. Bld-HrTH likely induces intracellular Ca2+ release followed by extracellular Ca2+ entry across the plasma membrane. Inositol-1,4,5-trisphosphate (InsP3) levels were greatly increased by Bld HrTH in a time- and dose-dependent manner, suggesting that InsP3 might be a Ca(2+)-mobilizing intracellular second messenger for Bld-HrTH.

Animals↗