Total Analysis and Purification of Cellular Proteins Binding to Cisplatin-Damaged DNA Using Submicron Beads

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• 作者：Takenori Tomohiro ; Jun-ichi Sawada ; Chika Sawa ; Hironori Nakura ; Shuhei Yoshida ; Masato Kodaka ; Mamoru Hatakeyama ; Haruma Kawaguchi ; Hiroshi Handa ; and Hiroaki Okuno
• 刊名：Bioconjugate Chemistry
• 出版年：2002
• 出版时间：March 2002
• 年：2002
• 卷：13
• 期：2
• 页码：163 - 166
• 全文大小：73K
• 年卷期：v.13,no.2(March 2002)
• ISSN：1520-4812

文摘

A high-performance affinity purification technique has been developed for cisplatin (CDDP)-damagedDNA binding proteins directly from crude nuclear extracts of HeLaS3 cell using novel submicronbeads synthesized by copolymerization of styrene and glycidyl methacrylate (GMA). The beadsdramatically decreased both nonspecific protein adsorption on solid surfaces and elution volume andsimplified the handling procedure. Preparation of the beads for purification was carried out byimmobilization of telomeric repeats, (TTAGGG)_n, on the surface after the reaction with CDDP. Atleast nine proteins clearly showed higher affinity to CDDP-DNA and were identified by amino acidsequence analysis including HMGB (high mobility group), hUBF (human upstream binding factor),and Ku autoantigen, which were previously reported to be components of CDDP-damaged DNA bindingproteins.