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A post-labeling technique for the iodination of DNA damage recognition proteins.
Billings, P C; Cryer, J E; Moy, L Y; Engelsberg, B N.
Afiliação
  • Billings PC; Department of Radiation Oncology, University of Pennsylvania School of Medicine, Philadelphia 19104, USA.
Cancer Biochem Biophys ; 14(4): 223-30, 1995 Jan.
Article em En | MEDLINE | ID: mdl-7767896
ABSTRACT
Major advances have recently been made in understanding the nucleotide excision repair pathway in mammalian cells. Although the signaling events responsible for initiating this process are not known, they probably involve proteins, i.e., damage recognition proteins (DRPs), which detect specific types of DNA damage. In this report, we describe a technique for labeling DNA damage recognition proteins. The procedure utilizes iodogen to radio-iodinate proteins bound to DNA modified with the cancer chemotherapy drug, cisplatin. Following iodination, bound proteins are eluted and analyzed on SDS-polyacrylamide gels. We have optimized this procedure such that the labeling reactions are rapid and employ small amounts of 125I. Using this procedure, we demonstrate that proteins of 28 and 40 kDa in MCF7 human breast epithelial cells bind to CDDP-DNA. This technique is sensitive and potentially will facilitate the identification of DRPs in samples containing limited amounts of protein, such as small tissue biopsy specimens obtained from patients undergoing diagnostic and/or therapeutic treatment.
Assuntos
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Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Dano ao DNA / Proteínas de Ligação a DNA Limite: Animals / Female / Humans Idioma: En Revista: Cancer Biochem Biophys Ano de publicação: 1995 Tipo de documento: Article País de afiliação: Estados Unidos
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Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Dano ao DNA / Proteínas de Ligação a DNA Limite: Animals / Female / Humans Idioma: En Revista: Cancer Biochem Biophys Ano de publicação: 1995 Tipo de documento: Article País de afiliação: Estados Unidos