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Identification of diphtheria toxin receptor and a nonproteinous diphtheria toxin-binding molecule in Vero cell membrane

Two substances possessing the ability to bind to diphtheria toxin (DT) were found to be present in a membrane fraction from DT-sensitive Vero cells. One of these substances was found on the basis of its ability to bind DT and inhibit its cytotoxic effect. This inhibitory substance competitively inhi...

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Formato: Texto
Lenguaje:English
Publicado: The Rockefeller University Press 1988
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Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2115223/
https://www.ncbi.nlm.nih.gov/pubmed/3417759
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description Two substances possessing the ability to bind to diphtheria toxin (DT) were found to be present in a membrane fraction from DT-sensitive Vero cells. One of these substances was found on the basis of its ability to bind DT and inhibit its cytotoxic effect. This inhibitory substance competitively inhibited the binding of DT to Vero cells. However this inhibitor could not bind to CRM197, the product of a missense mutation in the DT gene, and did not inhibit the binding of CRM197 to Vero cells. Moreover, similar levels of the inhibitory activity were observed in membrane fractions from DT-insensitive mouse cells, suggesting the inhibitor is not the DT receptor which is specifically present in DT-sensitive cells. The second DT-binding substance was found in the same Vero cell membrane preparation by assaying the binding of 125I-labeled CRM197. Such DT-binding activity could not be observed in membrane preparation from mouse L cells. From competition studies using labeled DT and CRM proteins, we conclude that this binding activity is due to the surface receptor for DT. Treatment of these substances with several enzymes revealed that the inhibitor was sensitive to certain RNases but resistant to proteases, whereas the DT receptor was resistant to RNase but sensitive to proteases. The receptor was solubilized and partially purified by chromatography on CM- Sepharose column. Immunoprecipitation and Western blotting analysis of the partially purified receptor revealed that a 14.5-kD protein is the DT receptor, or at least a component of it.
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spelling pubmed-21152232008-05-01 Identification of diphtheria toxin receptor and a nonproteinous diphtheria toxin-binding molecule in Vero cell membrane J Cell Biol Articles Two substances possessing the ability to bind to diphtheria toxin (DT) were found to be present in a membrane fraction from DT-sensitive Vero cells. One of these substances was found on the basis of its ability to bind DT and inhibit its cytotoxic effect. This inhibitory substance competitively inhibited the binding of DT to Vero cells. However this inhibitor could not bind to CRM197, the product of a missense mutation in the DT gene, and did not inhibit the binding of CRM197 to Vero cells. Moreover, similar levels of the inhibitory activity were observed in membrane fractions from DT-insensitive mouse cells, suggesting the inhibitor is not the DT receptor which is specifically present in DT-sensitive cells. The second DT-binding substance was found in the same Vero cell membrane preparation by assaying the binding of 125I-labeled CRM197. Such DT-binding activity could not be observed in membrane preparation from mouse L cells. From competition studies using labeled DT and CRM proteins, we conclude that this binding activity is due to the surface receptor for DT. Treatment of these substances with several enzymes revealed that the inhibitor was sensitive to certain RNases but resistant to proteases, whereas the DT receptor was resistant to RNase but sensitive to proteases. The receptor was solubilized and partially purified by chromatography on CM- Sepharose column. Immunoprecipitation and Western blotting analysis of the partially purified receptor revealed that a 14.5-kD protein is the DT receptor, or at least a component of it. The Rockefeller University Press 1988-08-01 /pmc/articles/PMC2115223/ /pubmed/3417759 Text en This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 4.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/4.0/).
spellingShingle Articles
Identification of diphtheria toxin receptor and a nonproteinous diphtheria toxin-binding molecule in Vero cell membrane
title Identification of diphtheria toxin receptor and a nonproteinous diphtheria toxin-binding molecule in Vero cell membrane
title_full Identification of diphtheria toxin receptor and a nonproteinous diphtheria toxin-binding molecule in Vero cell membrane
title_fullStr Identification of diphtheria toxin receptor and a nonproteinous diphtheria toxin-binding molecule in Vero cell membrane
title_full_unstemmed Identification of diphtheria toxin receptor and a nonproteinous diphtheria toxin-binding molecule in Vero cell membrane
title_short Identification of diphtheria toxin receptor and a nonproteinous diphtheria toxin-binding molecule in Vero cell membrane
title_sort identification of diphtheria toxin receptor and a nonproteinous diphtheria toxin-binding molecule in vero cell membrane
topic Articles
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2115223/
https://www.ncbi.nlm.nih.gov/pubmed/3417759