Plasma iron transport during egg laying and after oestrogen administration in the domestic fowl (Gallus domesticus). (1/62)

The concentrations of 59Fe and of radioiodinated transferrin and albumin were measured in the blood, liver, spleen, bone marrow and ova at different times after the injection of transferrin-bound 59Fe and the labelled proteins into non-laying, laying and oestrogen treated chickens. In the egg-laying and oestrogen-treated birds the 59Fe of the plasma was rapidly transferred from transferrin to another component with the properties of th phosphoprotein, phosvitin. Radioactive iron, and labelled transferrin and albumin to a lesser extent, entered the ova only while they were in the ovary. Relatively more labelled transferrin than albumin was found in all the tissues studied except in the ova, in which the two labelled proteins were present in the same relative concentration as in the plasma. It is concluded that, during egg laying and after oestrogen treatment, plasma iron bound to transferrin is taken up by the liver, incorporated into phosvitin and is then secreted into the plasma leading to elevation of the plasma iron concentration and transfer of iron to the ova.  (+info)

Phosphopeptides derived from hen egg yolk phosvitin: effect of molecular size on the calcium-binding properties. (2/62)

Two different phosphopeptide (PPP) fragments derived from partially dephosphorylated hen egg yolk phosvitin were prepared by tryptic digestion, and their Ca2+ binding property compared with that of commercial casein phosphopeptides (CPP). The smaller fragment of less than 1 kDa and O-phospho-1-serine did not bind Ca2+ to any significant extend, while PPP of 1-3 kDa showed a higher ability than CPP to render soluble calcium. The results show that not only the phosphoserine residues are critical for Ca2+ binding, but also the molecular size of the phosphopeptides.  (+info)

Purification and characterization of a casein kinase 2-type protein kinase from pea nuclei. (3/62)

Almost all the polyamine-stimulated protein kinase activity associated with the chromatin fraction of nuclei purified from etiolated pea (Pisum sativum L.) plumules is present in a single enzyme that can be extracted from chromatin by 0.35 molar NaCl. This protein kinase can be further purified over 2000-fold by salt fractionation and anion-exchange and casein-agarose column chromatography, after which it is more than 90% pure. The purified kinase has a specific activity of about 650 nanomoles per minute per milligram protein in the absence of polyamines, with either ATP or GTP as phosphoryl donor. Spermidine can stimulate its activity fourfold, with half-maximal activation at about 2 millimolar. Spermine and putrescine also stimulate activity, although somewhat less effectively. This kinase has a tetrameric alpha 2 beta 2 structure with a native molecular weight of 130,000, and subunit molecular weights of 36,000 for the catalytic subunit (alpha) and 29,000 for the regulatory subunit (beta). In western blot analyses, only the alpha subunit reacts strongly with polyclonal antibodies to a Drosophila casein kinase II. The pea kinase can use casein and phosvitin as artificial substrates, phosphorylating both the serine and threonine residues of casein. It has a pH optimum near 8.0, a Vmax of 1.5 micromoles per minute per milligram protein, and a Km for ATP of approximately 75 micromolar. Its activity can be almost completely inhibited by heparin at 5 micrograms per milliliter, but is relatively insensitive to concentrations of staurosporine, K252a, and chlorpromazine that strongly antagonize Ca(2+) -regulated protein kinases. These results are discussed in relation to recent findings that casein kinase 2-type kinases may phosphorylate trans-acting factors that bind to light-regulated promoters in plants.  (+info)

Spermine stimulation of a nuclear NII kinase from pea plumules and its role in the phosphorylation of a nuclear polypeptide. (4/62)

We have previously demonstrated that spermine stimulates the phosphorylation of a 47 kilodalton nuclear polypeptide from pea plumules (N Datta, LK Hardison, SJ Roux 1986 Plant Physiol 82: 681-684). In this paper we report that spermine stimulates the activity of a cyclic AMP independent casein kinase, partially purified from a chromatin fraction of pea plumule nuclei. This effect of spermine was substrate specific; i.e. with casein as substrate, spermine stimulated the kinase activity, and with phosvitin as substrate, spermine completely inhibited the activity. The stimulation by spermine of the casein kinase was, in part, due to the lowering of the Mg2+ requirement of the kinase. Heparin could partially inhibit this casein kinase activity and spermine completely overcame this inhibition. By further purification of the casein kinase extract on high performance liquid chromatography, we fractionated it into an NI and an NII kinase. Spermine stimulated the NII kinase by 5- to 6-fold but had no effect on the NI kinase. Using [gamma-32P]GTP, we have shown that spermine promotes the phosphorylation of the 47 kilodalton polypeptide(s) in isolated nuclei, at least in part by stimulating an NII kinase.  (+info)

The 35 kDa acid metallophosphatase of the frog Rana esculenta liver: studies on its cellular localization and protein phosphatase activity. (5/62)

The cellular localization of the 35 kDa, low molecular mass acid metallophosphatase (LMW AcPase) from the frog (Rana esculenta) liver and its activity towards P-Ser and P-Tyr phosphorylated peptides were studied. This enzyme was localized to the cytoplasm of hepatocytes but did not appear in other cells of liver tissue (endothelium, macrophages, blood cells). This LMW AcPase does not display activity towards (32)P-phosphorylase a under conditions standard for the enzymes of PPP family. Proteins containing P-Ser: rabbit (32)P-phosphorylasea and phosvitin are hydrolysed only at acidic pH and are poor substrates for this enzyme. The frog AcPase is not inhibited by okadaic acid and F(-) ions, the Ser/Thr protein phosphatase inhibitors. Moreover, the frog enzyme does not cross-react with specific antisera directed against N-terminal fragment of human PP2A and C-terminal conserved fragment of the eukaryotic PP2A catalytic subunits. These results exclude LMW AcPase from belonging to Ser/Thr protein phosphatases: PP1c or PP2Ac. In addition to P-Tyr, this enzyme hydrolyses efficiently at acidic pH P-Tyr phosphorylated peptides (hirudin and gastrin fragments). K(m) value for the hirudin fragment (7.55 +/- 1.59 x 10(-6) M) is 2-3 orders of magnitude lower in comparison with other substrates tested. The enzyme is inhibited competitively by typical inhibitors of protein tyrosine phosphatases (PTPases): sodium orthovanadate, molybdate and tungstate. These results may suggest that the LMW AcPase of frog liver can act as PTPase in vivo. A different cellular localization and different response to inhibition by tetrahedral oxyanions (molybdate, vanadate and tungstate) provide further evidence that LMW AcPase of frog liver is distinct from the mammalian tartrate-resistant acid phosphatases.  (+info)

A purified S6 kinase kinase from Xenopus eggs activates S6 kinase II and autophosphorylates on serine, threonine, and tyrosine residues. (6/62)

S6 kinases I and II have been purified previously from Xenopus eggs and shown to be activated by phosphorylation on serine and threonine residues. An S6 kinase clone, closely related to S6 kinase II, was subsequently identified and the protein product was expressed in a baculovirus system. Using this protein, termed "rsk" for Ribosomal Protein S6 Kinase, as a substrate, we have purified to homogeneity from unfertilized Xenopus eggs a 41-kDa serine/threonine kinase termed rsk kinase. Both microtubule-associated protein-2 and myelin basic protein are good substrates for rsk kinase, whereas alpha-casein, histone H1, protamine, and phosvitin are not. rsk kinase is inhibited by low concentrations of heparin as well as by beta-glycerophosphate and calcium. Activation of rsk kinase during Xenopus oocyte maturation is correlated with phosphorylation on threonine and tyrosine residues. However, in vitro, rsk kinase undergoes autophosphorylation on serine, threonine, and tyrosine residues, identifying it as a "dual specificity" enzyme. Purified rsk kinase can be inactivated in vitro by either a 37-kDa T-cell protein-tyrosine phosphatase or the serine/threonine protein phosphatase 2A. Phosphatase-treated S6KII can be reactivated by rsk kinase, and S6 kinase activity in resting oocyte extracts increases significantly when purified rsk kinase is added. The availability of purified rsk kinase will enhance study of the signal transduction pathway(s) regulating phosphorylation of ribosomal protein S6 in Xenopus oocytes.  (+info)

A novel nuclear 42-kDa casein kinase identified in Chironomus tentans. (7/62)

We have purified and characterised an apparently novel nuclear 42-kDa casein kinase from epithelial cells of Chironomus tentans which comigrates with a phosphoprotein associated with transcriptionally active salivary gland genes. The protein kinase promotes phosphorylation of casein and phosvitin, using either ATP or GTP as phosphate donors, and undergoes autophosphorylation. The casein kinase activity of the 42-kDa protein is sensitive to heparin, 5,6-dichloro-1-beta-D-ribofuranosylbezimidazole (DRB), spermine and spermidine indicating that it is a novel enzyme with similar but not identical properties to casein kinase II or nuclear protein kinase NII.  (+info)

Induction of a novel protein kinase in pupae of the silkworm Bombyx mori after infection with nuclear polyhedrosis virus. (8/62)

Protein kinases induced by Bombyx mori nuclear polyhedrosis virus in pupae of the silkworm B. mori were examined by activity gel analysis using phosvitin as a protein substrate. The method involved PAGE of the soluble fraction from pupae under native conditions and in the presence of SDS, followed by in situ renaturation of proteins and recovery of protein kinase activity in the intact gel. A novel protein kinase able to phosphorylate phosvitin was detected in the infected pupae from 2 days post-infection. This enzyme was not present in uninfected silkworms at any stage of the pupal period. The novel kinase activity was found by SDS-PAGE to be associated with a single polypeptide with an apparent M(r) of 50K. However, on electrophoresis under native conditions its activity was associated with a set of polypeptides with similar but not identical electrophoretic mobilities. Microheterogeneity of the catalytically active polypeptides suggests that the virus-induced protein kinase undergoes post-translational modification during the course of infection.  (+info)

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