Search PubMed⌕ Search

Biomedical subjects

K C Zoon

Publications and source records attributed to K C Zoon.

At least 55 records · Page 3Linked to original sources
PubMed1984

Microinjection of anti-interferon antibodies into cells does not inhibit the induction of an antiviral state by interferon.

Affinity-purified polyclonal antibodies directed against human lymphoblastoid interferon (IFN), Escherichia coli-derived human IFN-alpha 2, or two synthetic fragments of human IFN-alpha 1 all neutralized the antiviral activity of human alpha IFNs when added to the culture medium of MDBK cells together with IFNs. However, when these antibodies were microinjected into the cytoplasm or the nucleus of cells, subsequent treatment of the cells with IFNs induced full protection against vesicular stomatitis virus. This suggests that IFNs themselves need not act in the cytoplasmic compartment or the nucleus to induce an antiviral state.

Animals↗
PubMed1983

Human interferon alpha enters cells by receptor-mediated endocytosis.

A small number of Escherichia coli-derived human interferon alpha A molecules (up to approximately 800/cell) bind specifically to high-affinity cell surface receptors on bovine kidney cells at 4 degrees. When the cells are subsequently warmed to 37 degrees, the amount of surface-bound interferon alpha (IFN-alpha) as recognized by a radiolabeled monoclonal antibody rapidly decreases. Using 125I-IFN-alpha and treatment of cells with acetic acid/sodium chloride to remove surface-bound IFN, a similar decrease of surface-bound IFN and a corresponding increase in intracellular radiolabeled material is observed following the temperature shift. An analysis of the trichloroacetic acid precipitability of the radiolabeled material in the medium, removed by acid treatment or internalized, shows that IFN is degraded intracellularly and that its degradation products are then released into the medium. The process of uptake, degradation, and release of degraded material can be inhibited by the lysosomotropic agent chloroquine. A stable conjugate of IFN with 5-nm colloidal gold was prepared without a detectable loss of antiviral activity. As shown by transmission electron microscopy, the conjugate, bound to cells at 4 degrees, was found in clathrin-coated pits and later in receptosomes following a temperature shift to 37 degrees. Morphometric quantitation showed that an excess of native IFN added during binding of the conjugate in the cold reduced the appearance of conjugate in receptosomes by 80%. These studies demonstrate that at least a portion of receptor-bound IFN enters cells by receptor-mediated endocytosis.

Animals↗
PubMed1983

Orientation of a human leukocyte interferon molecule on its cell surface receptor: carboxyl terminus remains accessible to a monoclonal antibody made against a synthetic interferon fragment.

An 125I-labeled monoclonal antibody made against a synthetic 56-residue fragment of human leukocyte interferon (IFN) alpha 1 recognizes human, Escherichia coli-derived IFN alpha A bound to the surface of Madin-Darby bovine kidney cells. A major fraction of the antibody recognizes IFN specifically bound to the cells, because the number of bound antibody molecules corresponds to the number of cell-bound IFN molecules (as measured with radiolabeled ligand) and because the fraction of the IFN unspecifically bound to the cells is less than 10% of the total bound IFN. A synthetic carboxyl-terminal 16-residue IFN peptide, though not inhibiting binding of IFN to cells, inhibits binding of antibody to IFN. A recombinant IFN alpha A molecule with a carboxyl-terminal 13-residue deletion, though still able to compete for binding of IFN to cells, is not recognized by the antibody. Scatchard plot analysis of the binding data revealed apparent dissociation constants of 6.0 x 10(-10) M for the antibody-IFN interaction and of 4.0 x 10(-11) M for the IFN-cell receptor interaction. The antibody inhibits the binding of IFN to cells only weakly and neutralizes the antiviral activity of the ligand only when in a large molar excess. We conclude that the carboxyl-terminal 10-16 residues that are predicted from the cloned IFN cDNAs and that are present in some natural IFNs are not involved in binding to cells but are antigenic and hence exposed on the molecules' surface. That the carboxyl terminus is not directly involved in binding to cells is consistent with the observation that some IFNs with carboxyl-terminal deletions are biologically active.

Animals↗
PubMed1982

The purification and characterization of interferons.

Significant advances have been made in the last four years on the purification of interferons. Components of several human and mouse interferons have been purified to homogeneity. Partial characterization of these molecules has been achieved (i.e. amino acid composition, analytical maps of radiolabelled tryptic peptides, and amino terminal amino acid sequence). In addition, preliminary evidence has been accumulated which implies that the carbohydrate portion of the molecules may play a significant role in species specificity of interferons. At present, with minute quantities of homogenous interferons available, limited structural information has been obtained. Using current production and purification schemes, sufficient material can be obtained for further amino acid sequence studies and examination of the carbohydrate portion of the molecules. Additional structural characterization of the interferon gene(s). Furthermore, as sequence homologies have been observed between human and mouse interferons, all new structural information will help to establish the interrelationships in the family of interferon proteins.

Amino Acid Sequence↗
PubMed1979

Purification and partial characterization of human lymphoblast interferon.

One component of human lymphoblastoid interferon obtained from Namalwa cultures induced by Newcastle disease virus has been purified to a specific activity of 2.5 x 10(8) interferon units per mg of protein (protein content based on amino acid analysis). A single polypeptide species with an apparent molecular weight of 18,500 comigrating with the antiviral activity was observed by sodium dodecyl sulfate/polyacrylamide gel electrophoresis. Preliminary amino-terminal sequencing results support the conclusion that the interferon species is essentially homogeneous.

Amino Acids↗
PubMed1979

Large-scale production and concentration of human lymphoid interferon.

A stable and predictable production system is described for pilot plant quantities (milligram) of human lymphoid interferon, using suspension culture of an African Burkitt's lymphoma derived cell line Namalva with induction by Newcastle disease virus, B-1 strain. Cell cultures were grown in impeller-driven 50-liter fermentors with dilution of the postinduction culture using serum-free medium. High levels of dissolved oxygen were necessary for optimum cell growth. A total of 4,207 liters of interferon culture was produced in a series of 116 fermentor runs. An average yield of 3.5 log(10) international units of interferon per ml was realized before processing. Trichloroacetic acid was used to precipitate the interferon. An average of 3.35 log(10) international units of interferon per ml was recovered in the final nonpurified product.

Burkitt Lymphoma↗
PubMed1978

Production of human lymphoblastoid interferon by Namalva cells.

Optimum conditions for growth and interferon production by a human lymphoblastoid cell line, Namalva, have been studied. Adaptation to large-scale production is possible utilizing either Sendai virus or Newcastle disease virus. Priming of cultures before induction is unnecessary. The interferon produced has properties similar to human leukocyte interferon. The production of lymphoblastoid interferon per cell is increased two- to fourfold after dilution with serum-free medium of a saturation-density culture of Namalva induced with Newcastle disease virus. Maximum interferon yields were obtained 27 h after the addition of virus, using cultures diluted to 4 X 10(5) to 9 X 10(5) cells per ml. The presence of glutamine in the dilution medium was required for maximum interferon production. Newcastle disease virus appeared to inhibit the rates of RNA and protein synthesis more effectively in the diluted cultures.

Animals↗
PubMed1977

Human lymphoblastoid interferon. Large scale production and partial purification.

Human lymphoblastoid interferon was produced on an 800-liter scale (2.6 X 10(9) units) by induction of Namalva cells with Newcastle disease virus, strain B1. The interferon was partially purified by anti-leukocyte interferon affinity chromatography, sulfopropyl Sephadex ion exchange chromatography, isoelectric focusing, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Recovery of interferon after gel electrophoresis varied from 11 to 33% based on the original crude material, with about 35,000-fold purification. The gel electrophoresis resolved the antiviral activity into two components with apparent molecular weights of 18,000 and 22,000; treatment with glycosidases resulted in all the activity being associated with the lower molecular weight species. Interferon activity could be completely (85 to 113%) recovered from the gels by elution into a buffer containing sodium dodecyl sulfate. The presence of sodium dodecyl sulfate did not appear to affect the assay of interferon. The protein could also be completely (75 to 106%) eluted from gels stained with coomassie blue, again with no loss in activity.

Cell Line↗