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J Morser

Publications and source records attributed to J Morser.

17 recordsLinked to original sources
PubMed1992

Structure-function studies of the epidermal growth factor domains of human thrombomodulin.

Structure-function relationships in the 6 epidermal growth factor-like domains of human thrombomodulin (TME, residues 227-462) were studied by deletion mutagenesis. Purified and characterised proteins were used for kinetic studies. Deletion of EGF1, EGF2 and residues 310-332 in EGF3 had no effect on thrombin binding (Kd) or on kcat/KM for protein C activation by the thrombin-thrombomodulin complex. Deletion of the rest of EGF3 and the interdomain loop between EGF3 and EGF4 had no effect on Kd but decreased kcat/KM to 10% of TME. Deletion of residues 447-462 of EGF6 had no effect on kcat/KM but increased Kd for thrombin approximately 6-fold. Thus, the region 333-350 in EGF3-4 is critical for protein C activation by the thrombin-thrombomodulin complex and the region 447-462 in EGF6 is critical for thrombin binding.

Base Sequence↗
PubMed1992

Oxidation of a specific methionine in thrombomodulin by activated neutrophil products blocks cofactor activity. A potential rapid mechanism for modulation of coagulation.

Endothelial thrombomodulin (TM) plays a critical role in hemostasis as a cofactor for thrombin-dependent formation of activated protein C, a potent anticoagulant. Chloramine T, H2O2, or hypochlorous acid generated from H2O2 by myeloperoxidase rapidly destroy 75-90% of TM cofactor activity. Activated PMN, the primary in vivo source of biological oxidants, also rapidly inactivate TM. Oxidation of TM by PMN is inhibited by diphenylene iodonium, an inhibitor of NADPH oxidase. Both Met291 and Met388 in the six epidermal growth factor-like repeat domain are oxidized; however, only substitutions of Met388 lead to TM analogues that resist oxidative inactivation. We suggest that in inflamed tissues activated PMN may inactivate TM and demonstrate further evidence of the interaction between the inflammatory process and induction of thrombotic potential.

Adult↗
PubMed1991

Intravenous recombinant soluble human thrombomodulin prevents venous thrombosis in a rat model.

Thrombomodulin is an endothelial surface thrombin receptor. Thrombin bound to thrombomodulin loses all procoagulant activity and instead activates the protein C anticoagulant pathway. We developed a recombinant thrombomodulin analog and compared the effects of recombinant thrombomodulin (100 micrograms/ea), saline (controls), recombinant hirudin (1.0 mg/kg), and heparin (100 units/kg) on thrombus formation, activated partial thromboplastin time, and tail transection bleeding time in a rat model of stasis-induced venous thrombosis. Results showed that thrombus was detected in the vena cava in six of the six rats treated with saline solution, in zero of the six rats treated with recombinant thrombomodulin (p less than 0.05), in one of six rats treated with recombinant hirudin (p less than 0.05), and in zero of six rats treated with heparin (p less than 0.05). The activated partial thromboplastin time in rats receiving recombinant thrombomodulin was slightly longer than controls (22 +/- 8 vs 37 +/- 6, p less than 0.05). The bleeding times in rats receiving recombinant thrombomodulin were approximately twice as long as controls (215 +/- 68 vs 545 +/- 173, p = 0.037). In all rats treated with recombinant hirudin or heparin, activated partial thromboplastin times were greater than 120 seconds and bleeding times were greater than 1200 seconds. We conclude that recombinant thrombomodulin inhibits venous thrombosis in a rat model with less prolongation of activated partial thromboplastin time and bleeding time than heparin or hirudin.

Animals↗
PubMed1990

Differential effect of platelets on plasminogen activation by tissue plasminogen activator, urokinase, and streptokinase.

In this report, we have examined the effects of platelets on plasminogen activation by different activators. Platelets enhance activation of plasminogen by both 1- and 2-chain tissue plasminogen activator (t-PA). The primary effect of platelets is to lower the Km with a corresponding 5-8-fold increase in the kcat/Km. The effect is saturable with respect to the platelet concentration. Platelets enhance activation of both glu- and lys-plasminogen by t-PA. Platelets have no effect on plasminogen activation by streptokinase, and high and low molecular weight urokinase. Thus, there are marked differences in the effects of platelets on plasminogen activation depending on the plasminogen activator. These differences are likely to reflect differences in the interaction between platelets and the plasminogen activators.

Blood Platelets↗
PubMed1979

Export of proteins from oocytes of Xenopus laevis.

When human lymphoblastoid mRNA was microinjected into X. laevis oocytes, titers of interferon rapidly reached a maximum inside the oocyte while accumulation of interferon continued in the incubation medium for at least 45 hr. If interferon protein was injected into oocytes it was rapidly inactivated. Significantly, newly synthesized interferon but not injected interferon was found to be membrane-associated. Further experiments involving the co-injection of mRNAs coding for secretory proteins (guinea pig milk proteins and human interferon) and nonsecretory proteins (rabbit globin) revealed that only the secretory proteins were exported from the oocyte. Moreover, different proteins were exported at different rates. A distinct subclass of newly synthesized oocyte proteins of unknown function also accumulated in the incubation medium. Since the information encoded in the messenger RNAs of secretory proteins is sufficient to specify synthesis, compartmentation and secretion of these proteins, the oocyte may provide a complete system for the analysis of the secretory process.

Animals↗
PubMed1979

The effect of inhibitors of glycosylation on interferon production in human lymphoblastoid cells.

Interferon production was inhibited in the Namalwa line of human lymphoblastoid cells by treatment with 2-deoxy-D-glucose or D-glucosamine. D-Glucosamine also inhibited protein synthesis and the cells were no longer viable, whereas 2-deoxy-D-glucose allowed protein and RNA synthesis to continue at control rates, and the cells remained fully viable. It is concluded that a glycosylation step is essential for production of lymphoblastoid interferon.

Burkitt Lymphoma↗
PubMed1979

The toxic effect of double-stranded RNA for interferon-treated cells: evidence for a heterogeneous cellular response and the role of the cell nucleus.

When L929 cells were treated with interferon and subsequently with poly(rI).poly(rC), there was a pronounced toxic effect. Most of the cells lysed, but some survived and grew at the same rate as control cells to yield cells which were as sensitive to the effects of interferon and poly(rI).poly(rC) as the original population. The proportion of surviving cells did not vary with either the cell cycle or the cell density. The treated cells produced interferon and some of the interferon was produced by the resistant cells. Cells which had been X-irradiated before treatment with interferon and poly(rI).poly(rC) behaved similarly so cell division was not necessary for the development of toxicity. The toxic effect also developed when cells were enucleated with the aid of cytochalas in B after treatment with interferon, but not if they were enucleated before treatment. It is concluded that the nucleus is essential for interferon to exert its effect on the cells, but not for the development of cytotoxicity after the addition of poly(rI).poly(rC).

Animals↗
PubMed1979

The toxic effect of double-stranded RNA for interferon-treated L cells: studies on membrane transport and with cell-free systems.

We have tested two possible explanations for the toxic effects observed in L cells treated with interferon followed by poly(rI).poly(rC), namely (1) that the toxicity was preceded by and due to a change in membrane permeability which could be monitored by measurement of ion fluxes, or (2) that the toxicity was due to the action of the inhibitors of protein synthesis which are known to be formed when extracts from interferon-treated cells are treated with poly(rI).poly(rC) in vitro. Neither explanation was found to be correct.

Animals↗
PubMed1979

Characterization of interferon messenger RNA from human lymphoblastoid cells.

After treatment with Sendai virus, Namalwa cells form large amounts of interferon. RNA extracted from treated whole cells or from their polysomes was injected into Xenopus laevis oocytes and the interferon formed was titrated. The results show that the amount of interferon mRNA was maximal by 9 h after treatment of the cells with Sendai virus and then declined. Sucrose gradient centrifugation of the mRNA gave substantial purification and showed that its size was 12 S.

Animals↗
PubMed1979

The effect of 5-bromodeoxyuridine on interferon production in human cells.

5-Bromodeoxyuridine (BrdUrd) increased interferon production by the Namalwa line of human lymphoblastoid cells treated with Sendai virus, but inhibited their growth. Thymidine, which also inhibited cell growth had no effect on interferon production, so that growth inhibition per se was not the cause of the stimulation. BrdUrd was incorporated into cellular DNA; 5-chlorodeoxyuridine and 5-iododeoxyuridine (which are also incorporated) increased the interferon yield, but 5-fluorodeoxyuridine (which is not incorporated) did not. Thymidine reduced both the incorporation of BrdUrd and its stimulatory effect on interferon production. Deoxycytidine (which prevents the cytotoxic effects of BrdUrd) had no effect on the stimulation. BrdUrd also stimulated interferon production in response to poly(rI) . poly(rC) in growing human diploid fibroblasts but not in SV40 virus-transformed human cells. Since BrdUrd was incorporated into the DNA of all these cells, we concluded that incorporation is necessary, but not sufficient for the stimulation of interferon formation.

Bromodeoxyuridine↗
PubMed1978

Differences in sialic acid content of human interferons.

Human leucocyte, lymphoblastoid and fibroblast interferons were separately treated with sialidase and the effect of this on their isoelectric focusing was examined using a system in which full dissociation of complexes occurred. Both leucocyte and lymphoblastoid interferons showed a single form with an isoelectric point which was unaltered by treatment with sialidase. In contrast, fibroblast interferon showed three forms which were reduced to one by treatment with the enzyme.

Fibroblasts↗
PubMed1976

Simultaneous translation of structural and nonstructural proteins from Semliki-forest-virus RNA in two eukaryotic systems in vitro.

The Semliki Forest virus genome, 42-S RNA, and the virus-specific intracellular 26-S RNA were translated in two cell-free protein-synthesising systems, the wheat germ extract, and a partially purified system from mammalian tissues. The 26-S RNA directed the synthesis of structual proteins only, as revealed by tryptic peptide mapping. About 75--80% of the radioactivity in the products comigrated with capsid and about 4--8% with envelope protein peptides. All the capsid peptides and the full-sized capsid protein were found in the products in vitro, no complete envelope protein was formed and fewer than half of the envelope peptides were detected. This result is consistent with reports that there is only one initiation site for the translation of virus structural proteins, and that the capsid protein is N-terminal in the polyprotein followed by envelope proteins. The systems programmed with 42-S RNA yielded virtually the same structural peptides. However, the bulk of the radioactivity was in peptides which did not comigrate with the structural ones. These peptides were mostly associated with relatively small-sized products. This shows that Semliki Forest virus 42-S RNA has at least two initiation sites, one for the structural proteins and the other(s) for the nonstructural proteins.

Cell Line↗
PubMed1976

Initiation of translation directed by 42S and 26S RNAs from Semliki Forest virus in vitro.

The proteins synthesized in vitro in response to 42S and 26S RNAs from Semliki Forest virus were labeled with formyl-[35S]methionine from initiator tRNA. One protein which comigrated with viral capsid protein was labeled under the direction of 26S RNA, and only one labeled peptide was detected after digestion with trypsin. Further digestion with pronase gave rise to the dipeptide fMet-AsN. Several labeled polypeptides were found in the 42S RNA directed product and these had molecular weights of up to 150,000. However, tryptic digestion of the product yielded only one formylmethionyl-labeled peptide, which had a different mobility from that directed by the 26S RNA. Further digestion with pronase gave a single dipeptide, fMet-Ala. This indicates that nonstructural proteins as large as 150,000 daltons are probably synthesized from one initiation site on the 42S RNA. Translation starting from the internal initiation site on the 42S RNA, which is equivalent to that on the 26S RNA, could not be detected under the conditions used. Internal initiation sites which are similarly inactive have also been detected in other viral RNAs (e.g., brome mosaic virus, tobacco mosaic virus, and polyoma 19S RNA) and this suggests that, although eukaryotic mRNAs can contain more than one initiation site for protein synthesis, only the site nearer the 5' terminus is active in vitro.

Cell-Free System↗
PubMed1975

Translation of Semliki forest virus 42S and 26S RNAs in a cell-free system derived from Escherichia coli.

The SFV 42S RNA and the intracellular 26S RNA have been translated in a prokaryotic cell-free system, the E. coli S30. About half of the [35S]methionine-labelled products directed by both RNAs had molecular weights larger than 20,000 on polyacrylamide gels. Both products contained tryptic peptides which comigrated with all the capsid and envelope protein-derived peptides. The most striking difference between the prokaryotic and eukaryotic systems lay in the translation of the 42 S RNA: The "42S RNA-specific nonstructural" peptides, which predominate in the eukaryotic systems, were apparently absent from the product translated by the prokaryotic system.

Cell-Free System↗
PubMed1975

Polysomes and initiation complexes in vitro induced by Semliki Forest virus 42S and 26S RNA.

Semliki Forest virus 42S and 26S RNA induce the formation of polysomes when translated in vitro in cell-free systems. After analysis on sucrose gradients the polysomal structures, containing prelabelled RNA and nascent peptide chains labelled with [35S]methionine, had sedimentation values from 100 to 200S in the case of 26S RNA and from 150 to over 250S with 42S RNA. After incubation in the presence of inhibitors of elongation 26S RNA was found in initiation complexes sedimenting at about 80S. Under identical conditions 42S RNA had a heterogenous sedimentation pattern, being attached to complexes sedimenting even faster than 250S. These structures, however, fulfilled the criteria of initiation complexes suggesting that there were more than one ribosome attached to each 42S RNA under conditions where 26S RNA bound only one.

Cell-Free System↗