Biomedical subjects

M Abdel-Ghany

Publications and source records attributed to M Abdel-Ghany.

26 records · Page 2Linked to original sources

PubMed1988

Specific dephosphorylation of phosphoproteins by protein-serine and -tyrosine kinases.

Five protein kinases are shown to serve as specific phosphatases in the absence of ADP. Although the rates of hydrolysis are very slow compared to the forward phosphorylation rates under optimal conditions, they are of the same order as the reverse reaction in the presence of ADP. Because cells contain approximately equal to 3 mM ATP, neither the reverse reaction nor the phosphatase is likely to play a physiological role. beta-casein B phosphorylated by the catalytic subunit of cAMP-dependent protein kinase (protein kinase A) is specifically dephosphorylated by protein kinase A but not by polypeptide-dependent protein kinase (protein kinase P). beta-casein B phosphorylated by protein kinase P is specifically dephosphorylated by protein kinase P but not by protein kinase A. Histone H1 phosphorylated by protein kinase C is dephosphorylated by the same enzyme in the absence of ADP. In all cases tested addition of ADP and F1-ATPase accelerates moderately the rate of dephosphorylation. Native H+-ATPase from yeast plasma membranes is isolated mainly in the phosphorylated form. It is dephosphorylated and rephosphorylated by protein kinase P but not by protein kinase A. Protein-tyrosine kinase of the epidermal growth factor receptor phosphorylates the random synthetic polypeptide poly(Glu80Tyr20). The phosphorylated polymer is specifically dephosphorylated in the absence of ADP by epidermal growth factor receptor preparations but not by insulin receptor preparations. The same polymer phosphorylated by insulin receptor is dephosphorylated by insulin receptor but not by epidermal growth factor receptor preparations. By using a cycle of dephosphorylation-rephosphorylation, it is possible to identify proteins that are phosphorylated by these protein kinases in vivo. Should this method be applicable to additional protein kinases, it should be possible to estimate the quantitative contribution of each protein kinase to a single phosphoprotein.

ErbB Receptors↗

PubMed1988

Phosphorylation of synthetic random polypeptides by protein kinase P and other protein-serine (threonine) kinases and stimulation or inhibition of kinase activities by microbial toxins.

A synthetic random polymer of threonine and glutamate (1:4.4) is readily phosphorylated by protein kinase P but not by five other protein-serine (threonine) kinases. A synthetic random polymer of serine and arginine (1:3) is readily phosphorylated by protein kinase A and protein kinase C but not by protein kinase P. Although the amino acid sequences surrounding the phosphorylated serine (threonine) residue have been demonstrated in studies with small synthetic polypeptides to be decisive factors in the rate at which they are phosphorylated, the findings with the large synthetic polypeptides suggest that in the case of proteins the size, the tertiary structure, and particularly the electrostatic interactions are equally or more important contributing factors. Syringomycin, a toxin from Pseudomonas syringae, and polymyxin B, from Bacillus polymyxa, stimulate protein kinase P, strongly inhibit protein kinase C, and have no effect on protein kinase A. Basic polypeptides with high lysine content are phosphorylated by ATP nonenzymatically.

Adenosine Triphosphate↗

PubMed1987

Effect of protein kinase P on phosphorylations catalyzed by the epidermal growth factor.

Protein kinase P (PK-P) activated by histones or certain other basic compounds has been purified previously from yeast [Yanagita, Y., Abdel-Ghany, M., Raden, D., Nelson, N. & Racker, E. (1987) Proc. Natl. Acad. Sci. USA 84, 925-929]. It is shown here that PK-P is present in solubilized membranes of A-431 carcinoma cells where it changes the epidermal growth factor (EGF) receptor kinase activity. Polylysine, a weak PK-P activator, inhibited the autophosphorylation of the EGF receptor both in the absence and presence of EGF. Increased PK-P activity induced by histone 1, a potent activator, gave rise to increased autophosphorylation of the EGF receptor as well as phosphorylation at tyrosine residues of numerous other endogenous membrane components. The stimulation by histone was particularly striking in the presence of EGF. A similar stimulation was achieved with polylysine and EGF on addition of yeast PK-P. However, addition of yeast PK-P in the presence of histone 1 markedly inhibited the EGF-stimulated phosphorylation of endogenous membrane proteins. We conclude from these results that the effect of PK-P on the EGF receptor takes place in three phases: at low levels PK-P inhibits the autophosphorylation, at intermediate levels it stimulates the autophosphorylation as well as the EGF-dependent phosphorylation of numerous other membrane proteins, and at high levels it inhibits the phosphorylation of these proteins.

Epidermal Growth Factor↗

PubMed1987

Polypeptide-dependent protein kinase from bakers' yeast.

The purification and properties of a protein serine kinase (PK-P) extracted with Triton X-100 from membranes of bakers' yeast are described. The enzyme is virtually inactive unless either a histone or a heat-stable polypeptide from yeast membranes and Mg2+ are added. Other divalent cations substitute for Mg2+ poorly or not at all; most of them, including Mn2+, inhibit when added in the presence of 5 mM Mg2+. The enzyme is unstable but can be stabilized by addition of 0.1% Triton X-100 and 20% glycerol. The final preparation shows, on silver-stained electrophoresis gels, two major bands (Mr 41,000 and 35,000). According to gel filtration the molecular weight of the active protein is about 75,000. Of the two subunits, only the smaller one appears to be autophosphorylated. In addition to casein, the enzyme phosphorylates several proteins including the H+-ATPase (Mr 100,000) in the yeast plasma membrane. In order to demonstrate the phosphorylation of the ATPase (up to 0.9 equivalents), exposure of the latter to an acid phosphatase was required. Other phosphorylated proteins include mRNA cap-binding protein from mammalian erythrocytes and yeast, a glucocorticoid receptor protein, and a preparation of the guanine nucleotide-binding proteins Gi and Go from brain. A partial purification of a natural activator from yeast plasma membranes is described.

Chemical Phenomena↗

PubMed1984

New protein kinase from plasma membrane of Ehrlich ascites tumor cells activated by natural polypeptides.

A polypeptide-dependent protein kinase was purified about 80-fold from an extract of plasma membranes of Ehrlich ascites tumor cells. The membranes were extracted with Nonidet P-40, and the extract was purified by ammonium sulfate fractionation and hydroxylapatite and affinity chromatography. The activity was stimulated 10-fold or more by polypeptide preparations from a variety of tissues, including placenta and hypothalamus. Polypeptide-dependent protein kinase had a pH optimum of about 7.5 and required Mg2+ for activity. Mn2+ at low concentrations (200 microM) stimulated enzyme activity somewhat but inhibited activity strongly at higher concentrations. The best available substrate for polypeptide-dependent protein kinase was beta-casein, and little or no phosphorylation was observed with alpha-casein, kappa-casein, phosvitin, alpha-lactalbumin, alpha-lactoglobulin, and histone. However, several endogenous substrates from plasma membranes of Ehrlich ascites tumor cells were phosphorylated. Polypeptide-dependent protein kinase activity was not inhibited by 10 mM N-ethylmaleimide, and this resistance was useful in differentiating this protein kinase from other protein kinases that were present in crude fractions and sensitive to the inhibitor.

Animals↗

PubMed1984

A placental polypeptide activator of a membranous protein kinase and its relation to histone 1.

Crude transforming growth factor preparations of placenta contain a polypeptide that is required for the activity of a protein kinase that has been purified from plasma membrane preparations of Ehrlich ascites tumor cells. The kinase activator has been separated from transforming growth factor beta by reversed-phase HPLC and affinity chromatography. Like the transforming growth factor, it is heat stable and trypsin labile, but it is not inactivated by dithiothreitol. In sodium dodecyl sulfate/polyacrylamide gel electrophoresis the purified preparation shows a major double band at about 31,000 daltons. Comparisons of electrophoretic mobility, protein kinase stimulatory activity, and cross-reactivity with an antibody against histone 1 suggest that the placental activator is identical with histone 1.

Cell Membrane↗

PubMed1984

Stimulation of glycolysis by placental polypeptides and inhibition by duramycin.

Placental polypeptides present in crude preparations of transforming growth factors stimulate glycolysis when added to quiescent 3T3 cells, normal rat kidney, and chick embryo fibroblasts. The stimulation was apparent over a time period of at least 90 min and was seen at glucose concentrations ranging from 1 to 30 mM. Duramycin, an antibiotic isolated from Streptomyces cinnamomeus, inhibits the polypeptide-stimulated and nonstimulated glycolysis of intact cells, since it permeabilizes cells to Pi and nucleotides. However, duramycin also inhibits the Na+-K+-ATPase as well as the ouabain-insensitive Mg2+-ATPase of plasma membranes. Duramycin has no effect on glycolysis catalyzed by cell-free extracts of Ehrlich ascites tumor cells in the presence of mitochondrial ATPase but partially inhibits glycolysis when ADP and Pi are generated by ATPases of plasma membrane preparations.

Adenosine Triphosphatases↗

PubMed1983

A membrane-bound human placental protein kinase activated by endogenous polypeptides.

A protein kinase (PPdPK) was purified from plasma membranes of human placenta. Phosphorylation of casein, but not of phosvitin or lactalbumin, by [gamma-32 P]ATP in the presence of PPdPK was stimulated about 10-fold by naturally occurring polypeptides prepared from a variety of sources similar to the procedure of Roberts et al. (Proc. Natl. Acad. Sci. U.S.A. 77, 3494-3498, 1980). The amino acid phosphorylated on casein was serine. According to gel exclusion chromatography the mol.wt. of PPdPK was 95 000. In autoradiograms, following polyacrylamide-gel electrophoresis, the autophosphorylation of PPdPK was greatly enhanced by the polypeptide activators.

Cell Compartmentation↗