Increased phosphate content of fibrinogen in vivo correlates with alteration in fibrinogen behaviour

Abstract To investigate the considerable variation in stimulation of serum aminotransferase activities by pyridoxal-5'-phosphate added in vitro, we determined the pyridoxal-5'-phosphate content of plasma, using the tyrosine decarboxylase reaction together with the catalytic activities of alanine aminotransferase and aspartate aminotransferase, with and without pyridoxal-5'-phosphate supplementation, within a group of normal human individuals. We found a very significant inverse linear relationship between plasma pyridoxal-5'-phosphate concentration and stimulation of the activities of these enzymes in serum after supplementation with pyridoxal-5'-phosphate. We conclude that the degree of stimulation of the apoenzyme of the two serum aminotransferases clearly depends on the pyridoxal-5'-phosphate concentration in vivo.

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Phosphorylation of rat thymus histones, its control and the effects thereon of γ-irradiation

Biochemical Journal ◽

10.1042/bj1620171 ◽

1977 ◽

Vol 162 (1) ◽

pp. 171-181 ◽

Cited By ~ 2

Author(s):

A Fónagy ◽

M G Ord ◽

L A Stocken

Keyword(s):

Nuclear Protein ◽

The Other ◽

Phosphate Content ◽

Γ Irradiation ◽

Histone Protein ◽

Histone Proteins ◽

32P Uptake ◽

Liver Nuclei

The phosphate content of rat thymus histones was determined. As expected for a replicating tissue, histones 1 and 2B were more phosphorylated and had higher 32P uptakes than did histones from resting liver nuclei; the other histones all showed 32P uptake, but the phosphate content and uptake of histone 2A was about half that for liver histone 2A. When thymus nuclei were incubated in a slightly hypo-osmotic medium, non-histone proteins and phosphorylated histones were released into solution; this was enhanced if ATP was present in the medium. [gamma-32P]ATP was incorporated into non-histone proteins, including protein P1, and into the ADP-ribosylated form of histone 1; negligible 32P was incprporated into the other, bound, histones. Histones 1 and 2B added to the incubation medium were extensively, and histones 2A and 4 slightly, phosphorylated. Histones released by increasing the ionic strength of the medium were phosphorylated. Added lysozyme and cytochrome c were neither bound nor phosphorylated, but added non-histone protein P1 was phosphorylated, causing other histones to be released from the nuclei, especially histones 2A and 3. The released histones were phosphorylated. gamma-Irradiation decreased 32P uptake into the non-ADP-ribosylated histones 1 and 4; phosphorylation of histone 1 in vitro was unaffected. The importance of non-histone proteins, ATP availability and nuclear protein kinases to the control of histone phosphorylation in vivo is discussed.

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Stimulation of postprandial in vivo glycogenesis and lipogenesis of rats fed high fructose diet with varied phosphate content

Nutrition Research ◽

10.1016/j.nutres.2009.12.004 ◽

2010 ◽

Vol 30 (2) ◽

pp. 151-155 ◽

Cited By ~ 5

Author(s):

Lama E. Mattar ◽

Melanie A. Mattar ◽

Malek Batal ◽

Youssef Mouneimne ◽

Omar A. Obeid

Keyword(s):

Phosphate Content ◽

High Fructose Diet ◽

High Fructose ◽

Stimulation Of

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Role of glycerol 3-phosphate and glycerophosphate acyltransferase in the nutritional control of hepatic triacylglycerol synthesis

Biochemical Journal ◽

10.1042/bj2040247 ◽

1982 ◽

Vol 204 (1) ◽

pp. 247-256 ◽

Cited By ~ 30

Author(s):

Peter E. Declercq ◽

Luc J. Debeer ◽

Guy P. Mannaerts

Keyword(s):

Liver Weight ◽

Isolated Hepatocytes ◽

Enzyme Analysis ◽

Marker Enzyme ◽

Phosphate Content ◽

Triacylglycerol Synthesis ◽

Acid Load ◽

Parenchymal Cells ◽

Hepatic Triacylglycerol

1. Glycerol 3-phosphate content of isolated hepatocytes from starved rats and of glycogen-depleted hepatocytes from fed rats was low and severely limited triacylglycerol synthesis. 2. Raising the glycerol 3-phosphate content by addition of precursors to the cells resulted in a hyperbolic-like relationship between triacylglycerol synthesis and cellular glycerol 3-phosphate content. Statistical analysis of the curves showed no significant differences between the nutritional states either at saturating or at subsaturating glycerol 3-phosphate content. 3. Vmax. of glycerophosphate acyltransferase measured in homogenized hepatocytes was decreased by 30–40% in starvation. There was no change in apparent Km for glycerol 3-phosphate. Since at saturating glycerol 3-phosphate content esterification rates in hepatocytes of both nutritional states were identical, the enzyme is not limiting esterification under this condition. 4. At subsaturating glycerol 3-phosphate content the flux through glycerophosphate acyltransferase necessarily limits esterification. Therefore one would expect a decrease in esterification in starvation under this condition. This was the case when triacylglycerol synthesis was plotted against intracellular glycerol 3-phosphate concentration, calculated from the cellular glycerol 3-phosphate content and the intracellular water space, which was smaller in hepatocytes from starved rats. 5. The data obtained in hepatocytes were extrapolated to the intact liver by using the number of parenchymal cells per g of liver as determined from marker-enzyme analysis and the liver weight per 100g body weight. The extrapolation suggested that glycerol 3-phosphate is limiting esterification in vivo for contents below 0.3–0.4 and 0.5–0.65μmol/g for livers from fed and starved animals respectively. Also for a given fatty acid load and a glycerol 3-phosphate content below 0.3μmol/g the liver may esterify less in the starved state. However, at the glycerol 3-phosphate contents measured in freeze-clamped livers (0.30 and 0.44μmol/g for the fed and starved state respectively), livers in both nutritional states seemed capable of esterifying similar amounts of fatty acids.

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Effect of glucagon on hepatic phenylalanine hydroxylase in vivo

Bioscience Reports ◽

10.1007/bf01116943 ◽

1985 ◽

Vol 5 (6) ◽

pp. 463-467 ◽

Cited By ~ 2

Author(s):

Edel Beirne ◽

Michael P. Carty ◽

John Donlon

Keyword(s):

Phenylalanine Hydroxylase ◽

High Protein Diet ◽

Physiological Changes ◽

Phosphate Content ◽

Hydroxylase Activity ◽

Human Phenylalanine Hydroxylase ◽

Stimulation Of ◽

Glucagon Concentration

Moderate doses of glucagon (20 μg/kg I.V.) are sufficient to stimulate rat hepatic phenylalanine hydroxylase in vivo. In addition, the stimulation of the tetrahydrobiopterin-dependent phenylalanine hydroxylase activity in livers of animals fed on a high-protein diet has been correlated with an elevated phosphate content. The tetrahydrobiopterin-dependent hydroxylase activity in these animals can be further elevated by glucagon-stimulated phosphorylation. These results indicate that physiological changes in glucagon concentration modulate rat liver phenylalanine hydroxylase activity in vivo. The current understanding of the role of phosphorylation in regulating human phenylalanine hydroxylase is also considered.

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Studies on the phosphorylation of protein kinase C-α

Biochemical Journal ◽

10.1042/bj2830515 ◽

1992 ◽

Vol 283 (2) ◽

pp. 515-518 ◽

Cited By ~ 67

Author(s):

C Pears ◽

S Stabel ◽

S Cazaubon ◽

P J Parker

Keyword(s):

Protein Kinase C ◽

Protein Kinase ◽

Alpha Activity ◽

Phosphate Content ◽

Phosphorylated Protein ◽

Kinase C ◽

Protein Kinase C Alpha ◽

The Relationship

A kinase-defective protein kinase C-alpha mutant is shown to be a phosphoprotein when expressed in COS-1 cells, indicating that intramolecular phosphorylation does not fully account for the phosphate content of protein kinase C-alpha. Furthermore, evidence is presented that the intermolecular phosphorylation of protein kinase C-alpha is due to an activity other than protein kinase C-alpha itself, and this phosphorylation appears to be necessary for protein kinase C-alpha activity. By contrast, the characteristic shift in apparent molecular mass consequent on phosphorylation in vivo can be accounted for by autophosphorylation, as demonstrated in vitro. The relationship between these phosphorylated protein kinase C-alpha species is discussed.

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The role of acetyl-CoA carboxylase phosphorylation in the control of mammary gland fatty acid synthesis during the starvation and re-feeding of lactating rats

Biochemical Journal ◽

10.1042/bj2370085 ◽

1986 ◽

Vol 237 (1) ◽

pp. 85-91 ◽

Cited By ~ 19

Author(s):

M R Munday ◽

D G Hardie

Keyword(s):

Mammary Gland ◽

Kinetic Parameters ◽

Protein Phosphatase ◽

Mammary Glands ◽

Phosphate Content ◽

Acetyl Coa Carboxylase ◽

Acetyl Coa ◽

Carboxylase Activity

Activation of acetyl-CoA carboxylase during incubation of crude extracts of lactating rat mammary gland with Mg2+ and citrate can be blocked by NaF, suggesting that it represents a dephosphorylation of the enzyme. The greater extent of activation in extracts from 24 h-starved rats (200%) compared with fed controls (70%) implies that the decrease in acetyl-CoA carboxylase activity in response to 24 h starvation may involve increased phosphorylation of the enzyme. Acetyl-CoA carboxylase was purified from the mammary glands of lactating rats in the presence of protein phosphatase inhibitors by avidin-Sepharose chromatography. Starvation of the rats for 24 h increased the concentration of citrate giving half-maximal activation by 75%, and decreased the Vmax. of the purified enzyme by 73%. This was associated with an increase in the alkali-labile phosphate content from 3.3 +/- 0.2 to 4.5 +/- 0.4 mol/mol of enzyme subunit. Starvation of lactating rats for 6 h, or short-term insulin deficiency induced by streptozotocin injection, did not effect the kinetic parameters or the phosphate content of acetyl-CoA carboxylase purified from mammary glands. The effects of 24 h starvation on the kinetic parameters and phosphate content of the purified enzyme were completely reversed by re-feeding for only 2.5 h. This effect was blocked if the animals were injected with streptozotocin before re-feeding, suggesting that the increase in plasma insulin that occurs on re-feeding was responsible for the activation of the enzyme. The effects of re-feeding 24 h-starved rats on the kinetic parameters and phosphate content of acetyl-CoA carboxylase could be mimicked by treating enzyme purified from 24 h-starved rats with protein phosphatase-2A in vitro. Our results suggest that, in mammary glands of 24 h-starved lactating rats, insulin brings about a dephosphorylation of acetyl-CoA carboxylase in vivo, which may be at least partly responsible for the reactivation of mammary lipogenesis in response to re-feeding.

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Potentiation of in vitro concentric work in mouse fast muscle

Journal of Applied Physiology ◽

10.1152/jappl.1998.84.1.236 ◽

1998 ◽

Vol 84 (1) ◽

pp. 236-243 ◽

Cited By ~ 36

Author(s):

R. W. Grange ◽

R. Vandenboom ◽

J. Xeni ◽

M. E. Houston

Keyword(s):

Extensor Digitorum Longus ◽

Conditioning Stimulus ◽

Muscle Length ◽

Extensor Digitorum Longus Muscle ◽

Extensor Digitorum ◽

Fast Muscle ◽

Phosphate Content ◽

Longus Muscle

Grange, R. W., R. Vandenboom, J. Xeni, and M. E. Houston.Potentiation of in vitro concentric work in mouse fast muscle. J. Appl. Physiol. 84(1): 236–243, 1998.—Phosphorylation of myosin regulatory light chain (R-LC) is associated with potentiated work and power during twitch afterloaded contractions in mouse extensor digitorum longus muscle [R. W. Grange, C. R. Cory, R. Vandenboom, and M. E. Houston. Am. J. Physiol. 269 ( Cell Physiol. 38): C713–C724, 1995]. We now describe the association between R-LC phosphorylation and potentiated concentric work when the extensor digitorum longus muscle is rhythmically shortened and lengthened to simulate contractions in vivo. Work output (at 25°C) was characterized at sine frequencies of 3, 5, 7, 10, and 15 Hz at excursions of 0.6, 1.2, and 1.6 mm (∼5, 9, and 13% optimal muscle length) at a low level of R-LC phosphorylation. Muscles stimulated during the sine function with a single twitch at specific times before or after the longest muscle length yielded maximal concentric work near the longest muscle length at a sine frequency of 7 Hz (e.g., excursion ∼9% optimal muscle length = 1.6 J/kg). Power increased linearly between sine frequencies of 3 and 15 Hz at all excursions (maximum ∼29 W). After a 5-Hz 20-s conditioning stimulus and coincident with a 3.7-fold increase in R-LC phosphate content (e.g., from 0.19 to 0.70 mol phosphate/mol R-LC), work at the three excursions and a sine frequency of 7 Hz was potentiated a mean of 25, 44, and 50% ( P < 0.05), respectively. The potentiated work during rhythmic contractions is consistent with enhanced interaction between actin and myosin in the force-generating states. On the basis of observations in skinned skeletal muscle fibers (H. L. Sweeney and J. T. Stull. Proc. Natl. Acad. Sci. USA 87: 414–418, 1990), this enhancement could result from increased phosphate incorporation by the myosin R-LC. Under the assumption that the predominant effect of the conditioning stimulus was to increase R-LC phosphate content, our data suggest that a similar mechanism may be evident in intact muscle.

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Changes in inorganic phosphate and force production in human skeletal muscle after cast immobilization

Journal of Applied Physiology ◽

10.1152/japplphysiol.00612.2004 ◽

2005 ◽

Vol 98 (1) ◽

pp. 307-314 ◽

Cited By ~ 25

Author(s):

Neeti Pathare ◽

Glenn A. Walter ◽

Jennifer E. Stevens ◽

Zhaohui Yang ◽

Enyi Okerke ◽

...

Keyword(s):

Skeletal Muscle ◽

Magnetic Resonance ◽

Inorganic Phosphate ◽

Force Production ◽

Resonance Spectroscopy ◽

Specific Force ◽

Phosphate Content ◽

Cast Immobilization ◽

Resting Energy

Cast immobilization is associated with decreases in muscle contractile area, specific force, and functional ability. The pathophysiological processes underlying the loss of specific force production as well as the role of metabolic alterations are not well understood. The aim of this study was to quantify changes in the resting energy-rich phosphate content and specific force production after immobilization.31P-magnetic resonance spectroscopy, three-dimensional magnetic resonance imaging, and isometric strength testing were performed in healthy subjects and patients with an ankle fracture after 7 wk of immobilization and during rehabilitation. Muscle biopsies were obtained in a subset of patients. After immobilization, there was a significant decrease in the specific plantar flexor torque and a significant increase in the inorganic phosphate (Pi) concentration ( P < 0.001) and the Pi-to-phosphocreatine (PCr) ratio ( P < 0.001). No significant change in the PCr content or basal pH was noted. During rehabilitation, both the Picontent and the Pi-to-PCr ratio decreased and specific torque increased, approaching control values after 10 wk of rehabilitation. Regression analysis showed an inverse relationship between the in vivo Piconcentration and specific torque ( r = 0.65, P < 0.01). In vitro force mechanics performed on skinned human muscle fibers demonstrated that varying the Pilevels within the ranges observed across individuals in vivo (4–10 mM) changed force production by ∼16%. In summary, our findings clearly depict a change in the resting energy-rich phosphate content of skeletal muscle with immobilization, which may negatively impact its force generation.

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Formation of n.m.r.-invisible ADP during renal ischaemia in rats

Biochemical Journal ◽

10.1042/bj2240241 ◽

1984 ◽

Vol 224 (1) ◽

pp. 241-246 ◽

Cited By ~ 32

Author(s):

M Stubbs ◽

D Freeman ◽

B D Ross

Keyword(s):

Inorganic Phosphate ◽

Metabolic Regulation ◽

Adenine Nucleotide ◽

Acid Extraction ◽

Phosphate Content ◽

Renal Ischaemia ◽

A Value ◽

Enzymic Assay

Measurement of the adenine nucleotide and inorganic phosphate content of normoxic and ischaemic kidney in vivo has been made, comparing enzymic assay (after freeze-clamping and acid extraction) with quantification by 31P-n.m.r. Both methods give similar results for ATP, and n.m.r. quantification of Pi gives a value 25-50% of that obtained by enzymic assay. ADP, which is largely invisible to n.m.r. in the normoxic kidney, remains invisible during ischaemia despite a 2-3 fold rise in enzymically assayed ADP. N.m.r. and enzymic assay of the acid extracts give similar values for all metabolites measured. The question of ADP binding in the kidney is discussed, as are the implications for the metabolic regulation of ADP-dependent reactions.

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