Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.21.4 (trypsin)
42,187 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Proteolytic digests of biologically active fractions of recombinant human leukocyte interferon A expressed in large quantities in Escherichia coli were analyzed by fast atom bombardment mass spectrometry and high-performance liquid chromatography. The values observed in the mass spectra of digests of the major fraction of recombinant human leukocyte interferon A with trypsin and Staphylococcus aureus protease V8 accounted for 93% of the amino acid sequences of human leukocyte interferon A predicted from the nucleotide sequence of the gene encoding the protein, indicating that the major fraction of recombinant human leukocyte interferon A was expressed with the same amino acid sequence as that translated from the nucleotide sequence of the gene encoding the protein. Mass spectrometry of proteolytic digests of two minor fractions of recombinant human leukocyte interferon A and mass and amino acid analyses of their high-performance liquid chromatography fractions showed that the amino group of the N-terminal amino acid residue of interferon was in part acetylated, and the Cys-1 and Cys-98 residues were oxidized to cysteic acid or linked to glutathione. These findings suggest that amino acid residues in recombinant proteins prepared in large quantities in E. coli are modified post-translationally.
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PMID:Chemical characterization of recombinant human leukocyte interferon A using fast atom bombardment mass spectrometry. 354 8

A mouse macrophage cytotoxic factor was purified to homogeneity from the serum-free culture supernatant of a mouse macrophage hybridoma clone, N/P-7-1, stimulated with lipopolysaccharide by gel filtration, affinity chromatography, anion-exchange chromatography, and polyacrylamide gel electrophoresis. The purified material was judged to be homogeneous as to the criteria of sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and has a relative molecular mass of 17,500, as determined by SDS-PAGE, or 55,000, as determined by gel filtration on columns of both Sephacryl S-200 and TSK G3000SW. It has an isoelectric point of 5.0, and is trypsin sensitive, stable at 56 degrees C and labile at pH less than 6. The cytotoxic activity of the purified factor could not be inhibited by various sugars and lectins. The production of the factor from N/P-7-1 triggered by macrophage-activating factor for cytotoxicity, but not by mouse recombinant gamma-interferon. The factor should be synthesized after lipopolysaccharide stimulation because treatment of N/P-7-1 cells with a metabolic inhibitor, emetine or actinomycin D, prevents the production.
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PMID:Purification and characterization of a cytotoxic factor produced by a mouse macrophage hybridoma. 383 93

Murine T cell replacing factor (TRF) was purified from a cellfree supernatant of a T cell hybridoma (B151K12) that constitutively produces TRF. Two assay systems for TRF activity were employed: 1) induction of anti-DNP IgG PFC responses in cultures of splenic B cells from DNP-KLH-primed BALB/c mice, and 2) induction of IgM PFC in chronic B cell leukemic cells (BCL1). The purification scheme consisted of ammonium sulfate precipitation, DEAE-cellulose chromatography, Blue-Sepharose chromatography, hydroxylapatite chromatography, gel permeation with fast protein liquid chromatography (FPLC), and disc polyacrylamide gel electrophoresis. Overall, TRF was purified approximately 34,000-fold with a maximum 3.8% recovery of activity, and the specific activity of the purified TRF was approximately 9.6 X 10(4) U/mg. The TRF that is active in these systems is distinct from the other lymphokines such as IL 1, IL 2, BCGFI (now known as BSFp1), and gamma-interferon. The TRF is extremely hydrophobic, with an apparent m.w. of 50,000 to 60,000 on gel permeation chromatography and 18,000 on SDS-PAGE under reducing conditions. Highly purified B151-TRF abrogated the activity by treatment with trypsin but not with RNase. Moreover, it bound to lima bean agglutinin-Sepharose specific for N-acetylgalactosamine residues, indicating that B151-TRF is a glycosylated glycoprotein containing N-acetylgalactosamine residues. The role of N-acetylgalactosamine residues on TRF activity was additionally substantiated by the fact that the addition of appropriate amounts of N-acetylgalactosamine in the assay systems for TRF preferentially induced a profound suppression for TRF-mediated PFC responses.
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PMID:Purification and physicochemical characterization of murine T cell replacing factor (TRF). 387 Nov 9

Cells of an antigen-specific T-cell clone, A37.4, were treated with the proteolytic enzymes trypsin or pronase to remove the T-cell antigen receptor. Removal of the receptor, analysed by surface labelling and non-reduced/reduced gel electrophoresis, stimulated the cells to synthesize new protein rapidly. New membrane protein was readily detectable within 3 h after reculture, and it was able to interact with antigen-presenting cells, since the T cells could be stimulated to produce interferon. Treatment of cells with metabolic inhibitors demonstrated that there was only a small cytoplasmic pool of protein, and de novo synthesis of mRNA was necessary for quantitative replacement of the membrane protein. The protein also required glycosylation for transport and insertion into the membrane. Despite rapid resynthesis of new receptor, when surface-labelled cells were recultured either with or without antigen-presenting cells, there was not a rapid turnover of the receptor. There was also no evidence of any loss of receptor from the membrane of antigen-stimulated cells or of any shedding of receptor into the culture medium.
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PMID:Synthesis and turnover of the putative T-lymphocyte antigen receptor. 387 43

A lymphokine inhibitory for cellular DNA synthesis (termed STIF) was isolated from the culture supernatants of concanavalin A (Con A)-stimulated SD rat spleen cells. STIF inhibited the DNA synthesis of mouse bone marrow cells as well as mouse leukemia cells. STIF has an apparent m.w. of 45,000 to 50,000 and is separable from IL 2, m.w. 20,000 to 25,000, by Sephacryl S-200 gel filtration, but not from immune interferon (IFN) having the same m.w. as STIF. Con A-Sepharose chromatography of the fraction containing STIF and IFN could separate these lymphokines into Con A-unbound and Con A-bound fractions, respectively. Further fractionation of the STIF fraction by DEAE-Sephadex A-50 or Mono Q-FPLC anion exchange chromatography indicated that the STIF fraction contained two components of STIF activity, both showing the same pI value (5.1 to 5.6) on flat-bed isoelectric focusing. STIF was characterized as a sugar-free lymphokine of trypsin-sensitive protein nature.
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PMID:Stimulated rat T cell-derived inhibitory factor for cellular DNA synthesis (STIF). I. Isolation and characterization. 391 68

Sera from C57Bl/6 mice treated orally with Ge-132 exhibited antitumour activity against Ehrlich (allogeneic) and RL male 1 (syngeneic) ascites tumours in BALB/c mice. Sera obtained from mice 24 h after Ge-132 administration displayed the greatest antitumour effect and this was dose dependent. Sera prepared from mice 12, 36, or 48 h after Ge-132 treatment had no protective effect. Circulating interferon (IFN) was induced at 24 h after administration of Ge-132 but was not detected in the sera at 12, 36, or 48 h after administration. The antiviral activity of sera from Ge-132-treated mice was inactivated by treatments with trypsin, low pH, and anti-IFN gamma antiserum. The inactivated preparations of serum IFN induced by Ge-132 did not exhibit antitumour activity when administered to tumour-bearing mice. These results suggest that antitumour activity in the sera of Ge-132-treated mice may be expressed through activities of Ge-132-induced lymphokine(s), such as IFN gamma.
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PMID:Ability of sera from mice treated with Ge-132, an organic germanium compound, to inhibit experimental murine ascites tumours. 393 36

Serum specimens from mice treated orally with Ge-132 (100 mg/kg) exhibited antitumor activity against Ehrlich (allogeneic) and RL 1 (syngeneic) ascites tumors in BALB/c mice. Sera obtained from mice 24 hours after Ge-132 administration displayed the highest antitumor effect and the antitumor activity was dose-dependent. Sera prepared from mice 12, 36 or 48 hours after Ge-132 treatment had no protective effect. Circulating interferon (IFN) was induced at 24 hours after administration. The antiviral activity of serum from Ge-132-treated mice was inactivated by treatment with trypsin, low pH, and anti-IFN-gamma antiserum. The inactivated preparations of serum IFN induced by Ge-132 did not show antitumor activity when administered to mice bearing Ehrlich ascites tumors. These results suggest that the antitumor activity in the sera of Ge-132-treated mice may have been expressed through IFN-gamma which was induced by Ge-132.
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PMID:[Ability of sera from mice treated with Ge-132, an organo-germanium compound, to inhibit experimental murine ascites tumors]. 393 51

A serum viral inhibitor (SVI) was isolated from goats and partially characterized. The inhibitor prevented the cytopathic effects of vesicular stomatitis virus, encephalomyocarditis virus, and a caprine herpesvirus, indicating broad antiviral activity. The SVI was distinct from interferon because SVI did not induce an antiviral state in cells (ie, lack of protection of SVI-treated cells from virus challenge). The SVI had activity on heterologous cells, including human, bovine, and ovine cells. The lack of antiviral activity in mouse cells indicated that SVI was not an antibody. Like fibroblast interferon, however, SVI was sensitive to trypsin, was acid stable at pH 2 and 4 C for 4 days, was heat stable at 56 C for 1 hour, and could not be sedimented by centrifugation at 100,000 X g for 4 hours.
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PMID:Preliminary characterization of a serum viral inhibitor in goats. 407 46

Buckler, Charles E. (National Institute of Allergy and Infectious Diseases, Bethesda, Md.), and Samuel Baron. Antiviral action of mouse interferon in heterologous cells. J. Bacteriol. 91:231-235. 1966.-The antiviral action of mouse interferon in cell cultures of mouse, hamster, rat, chicken, and monkey origin was investigated. Using a vesicular stomatitis virus (VSV) plaque reduction test, we found that mouse serum interferon, assayed on closely related rat or hamster cells, exerted 5% of its homologous antiviral activity. This activity was characterized as interferon by its temperature of inactivation, trypsin sensitivity, nonsedimentability, stability at pH 2, lack of inactivation by antibody to virus, and inability to be washed off cells. In the more distantly related chicken and monkey cells, mouse interferon had less than 0.1% of its homologous activity. Conflicting reports of heterologous activity of chicken and mouse interferon preparations may result in part from the observed action of noninterferon inhibitors of vaccinia virus. These inhibitors, like interferon, are stable at pH 2. They are present in mouse serum, mouse lung extracts, and allantoic fluid, and they prevent the development of vaccinia plaques when allowed to remain in contact with cells during virus growth. Unlike interferon the inhibitors are removed by adequate washing of cells prior to virus challenge, and they are not active in the VSV assay system. These findings reemphasize the need for thorough characterization of interferon preparations.
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PMID:Antiviral action of mouse interferon in heterologous cells. 428 34

The production of interferon by porcine kidney (PK(15)) cell culture in response to viral and synthetic inducers was studied. The inducers used included a synthetic double-stranded polyribonucleotide, polyriboinosinic-polyribocytidylic acid (Poly I:C), swine influenza virus and three strains of pseudorabies virus. Following exposure to these inducers cell culture fluids were examined for interferon by the plaque-reduction method. The Poly I:C and the swine influenza virus induced production of interferon by PK(15) cell cultures, whereas, all three strains of pseudorabies virus at the two concentrations tested failed to induce production of interferon in vitro. The antiviral substance produced in PK(15) cells was identified as an interferon because it was pH stable, non-dialyzable, sensitive to trypsin, non-sedimentable, relatively heat stable, host-species specific and it possessed broad-spectrum antiviral activity. The latter was demonstrated by inhibition of vesicular stomatitis, vaccinia and pseudorabies viruses. Differences in interferon activity against the different viruses were observed.
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PMID:Swine interferon. I. Induction in porcine cell cultures with viral and synthetic inducers. 434 Mar 45


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