Subcellular distribution of cyclic AMP‐dependent protein kinase activity and of cyclic AMP‐binding proteins in human platelets

Salama, Samih; Haslam, Richard J.

doi:10.1016/0014-5793(81)81127-1

Cited by 11 publications

(10 citation statements)

References 23 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The A2-type receptors also differ in their tissue distribution. A2AAR is highly expressed in the striatum, nucleus accumbens, and olfactory tubercle 66 ; it has also been demonstrated to be expressed in platelets 68, 69 , immune cells 70, 71 , lung 66 , heart 72 and the vasculature 66, 73 . The A2BAR is predominantly expressed in the vasculature, retina and brain, with lower levels of expression in various cells and tissues, including platelets at baseline 66, 74 .…”

Section: Adenosine Receptors: Classification and Distributionmentioning

confidence: 99%

A2 Adenosine Receptors and Vascular Pathologies

Johnston-Cox

Koupenova²,

Ravid³

2012

ATVB

View full text Add to dashboard Cite

Cardiovascular disease, a leading cause of death and morbidity, is regulated, among various factors, by inflammation. The level of the metabolite adenosine is augmented under stress, including inflammatory, hypoxic, or injurious events. Adenosine has been shown to affect various physiological and pathological processes, largely through 1 or more of its 4 types of receptors: the A1 and A3 adenylyl cyclase inhibitory receptors and the A2A and A2B adenylyl cyclase stimulatory receptors. This article focuses on reviewing common and distinct effects of the 2 A2-type adenosine receptors on vascular disease and the mechanisms involved. Understanding the pathogenesis of vascular disease mediated by these receptors is important to the development of therapeutics and to the prevention and management of disease.

show abstract

Section: Adenosine Receptors: Classification and Distributionmentioning

confidence: 99%

A2 Adenosine Receptors and Vascular Pathologies

Johnston-Cox

Koupenova²,

Ravid³

2012

ATVB

View full text Add to dashboard Cite

show abstract

“…Platelet particulate fractions were resuspended in the original volume of platelet lysis medium or in half the original volume of extraction buffers. In some experiments, platelets were disrupted by sonication, and supernatant and particulate fractions were prepared as described by Salama & Haslam (1981a).…”

Section: Methodsmentioning

confidence: 99%

“…The reaction was started by addition of 0.1 ml of column fraction or 0.05 ml of platelet fraction and the mixture was incubated for 3min at 30°C. In both assays, enzyme activity was terminated by addition of trichloroacetic acid, and the 32plabelled protein was isolated on glass-fibre filters, washed and counted for 32P radioactivity as described previously (Salama & Haslam, 1981a). One unit of protein kinase activity is defined as that transferring 1 pmol of [32P]phosphate from [y_32P]_ ATP to histone/min at 30°C.…”

Section: Methodsmentioning

confidence: 99%

“…Photoaffinity labelling ofcyclic AMP-binding proteins. This was performed with 0.5 uM-8-azidocyclic [32P]AMP, as described previously (Salama & Haslam, 1981a). After their separation by SDS/polyacrylamide-gel electrophoresis, 32plabelled polypeptides were detected by autoradiography and quantified by using a Joyce-Loebl microdensitometer.…”

Section: Methodsmentioning

confidence: 99%

“…Thus much more cyclic AMP-dependent protein kinase activity was found in the platelet particulate fraction when platelets were lysed in the presence rather than in the absence of inhibitors of Ca2 + -dependent proteinase (Salama & Haslam, 1981a). Two studies utilizing 8-azido-cyclic [32P]AMP as a photoaffinity label have shown that, when platelets were sonicated, the RI, regulatory subunits of platelet cyclic AMP-dependent protein kinase were found almost exclusively in the supernatant fraction, whereas at least half of the RI subunits were found in the particulate fraction (Lyons, 1980;Salama & Haslam, 1981a). However, a third study (Chambers et al, 1982) failed to detect RI, regulatory subunits in any subcellular fraction, though RI subunits were found in all platelet fractions.…”

mentioning

confidence: 98%

See 2 more Smart Citations

Characterization of the protein kinase activities of human platelet supernatant and particulate fractions

Salama¹,

Haslam²

1984

Biochemical Journal

View full text Add to dashboard Cite

After human platelets were lysed by freezing and thawing in the presence of EDTA, about 35% of the total cyclic AMP-dependent protein kinase activity was specifically associated with the particulate fraction. In contrast, Ca2+-activated phospholipid-dependent protein kinase was found exclusively in the soluble fraction. Photoaffinity labelling of the regulatory subunits of cyclic AMP-dependent protein kinase with 8-azido-cyclic [32P]AMP indicated that platelet lysate contained a 4-fold excess of 49 000-Da RI subunits over 55 000-Da RII subunits. The RI and RII subunits were found almost entirely in the particulate and soluble fractions respectively. Chromatography of the soluble fraction on DEAE-cellulose demonstrated a single peak of cyclic AMP-dependent activity with the elution characteristics and regulatory subunits characteristic of the type-II enzyme. A major enzyme peak containing Ca2+-activated phospholipid-dependent protein kinase was eluted before the type-II enzyme, but no type-I cyclic AMP-dependent activity was normally observed in the soluble fraction. The particulate cyclic AMP-dependent protein kinase and associated RI subunits were solubilized by buffers containing 0.1 or 0.5% (w/v) Triton X-100, but not by extraction with 0.5 M-NaCl, indicating that this enzyme is firmly membrane-bound, either as an integral membrane protein or via an anchor protein. DEAE-cellulose chromatography of the Triton X-100 extracts demonstrated the presence of both type-I cyclic AMP-dependent holoenzyme and free RI subunits. These results show that platelets contain three main protein kinase activities detectable with histone substrates, namely a membrane-bound type-I cyclic AMP-dependent enzyme, a soluble type-II cyclic AMP-dependent enzyme and Ca2+-activated phospholipid-dependent protein kinase, which was soluble in lysates containing EDTA.

show abstract

Protein kinases in normal human blood cells

et al. 1983

View full text Add to dashboard Cite

Protein kinases active on basic and acidic artificial substrates were investigated in normal human erythrocytes, platelets, polymorphonuclear and mononuclear cells. These two types of protein kinases were partially purified by affinity chromatography, then assayed for their enzymatic activity using [gamma-32P]ATP or GTP as phosphoryl donor. Partially purified kinases active on acidic substrates were subjected to high-performance liquid chromatography (HPLC). Protein kinases active on basic substrates were analyzed by cellulose acetate electrophoresis of crude cellular extracts and the influence of 3'5' cyclic AMP was studied. Three forms of casein-phosvitin kinases could be distinguished according to their molecular weight (165 K, 38 K, and 31 K). The 165 K species, in contrast to the light species, can use GTP instead of ATP as phosphoryl donor and corresponds to the "so-called" casein kinase 2. This form is very sensitive to proteolysis and, when partial purification is performed without the addition of various antiproteolytic agents, it is degraded into 120-135 K and 105-115 K active species; this artefactual degradative process is especially active in platelet extracts. As many as eight different active bands of histone and protamine kinases can be separated by cellulose acetate electrophoresis, several of them being stimulated by cyclic AMP. Isozymic patterns of protein kinases, levels of activity on the different substrates, and utilization of ATP and GTP were found to be specific for each cell type. These results suggest the possibility of using protein kinases as markers for cell differentiation.

show abstract

Subcellular distribution of cyclic AMP‐dependent protein kinase activity and of cyclic AMP‐binding proteins in human platelets

Cited by 11 publications

References 23 publications

A2 Adenosine Receptors and Vascular Pathologies

A2 Adenosine Receptors and Vascular Pathologies

Characterization of the protein kinase activities of human platelet supernatant and particulate fractions

Protein kinases in normal human blood cells

Contact Info

Product

Resources

About