2007 Fiscal Year Final Research Report Summary
DETERMINATION OF UNKNOWN POLY(ADP-RIBOSYL)ATED PROTEINS AND BINDING SITES
| Project/Area Number |
18590277
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
General medical chemistry
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| Research Institution | Nagahama Institute of Bio-Science and Technology |
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Principal Investigator |
MIWA Masanao Nagahama Institute of Bio-Science and Technology, FACULTY OF BIO-SCIENCE AND TECHNOLOGY, PROFESSOR (20012750)
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| Co-Investigator(Kenkyū-buntansha) |
KAMEMURA Kazuo NAGAHAMA INSTITUTE OF BIO-SCIENCE AND TECHNOLOGY, FACULTY OF BIO-SCIENCE AND TECHNOLOGY, LECTURER (00399437)
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| Project Period (FY) |
2006 – 2007
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| Keywords | posttranslational modification / polyADP-ribosylation / poly(ADP-ribose) gllycohydrolase / Drosophila melanogaster mutant / poly(ADP-ribose) nolymerase / antibody against poly(ADP-ribose) |
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| Research Abstract |
To understand the physiological function of polyADP-ribosylation, it is absolutely necessary to identify the acceptor proteins of polyADP-ribosylation. We prepared and quantities of monoclonal antibody against poly(ADP-ribose) and purified it for affinity chromatography. During the course of experiments, we noticed that the extracted polyADP-ribosylated proteins did not bind to the affinity column suggesting that there are other proteins binding to poly(ADP-ribose) that inhibit the binding of polyADP-ribosylated proteins to the column. To solve these problems, we used human kidney cell line (293T) as cell lysate and synthesized polyADP-ribosylated proteins in vitro and checked the ideal condition of extraction of polyADP-ribosylated proteins. It turned out that extraction buffer should contain 0.1% SDS for better extraction. For extracting the polyADP-ribosylated proteins from Drosophila mutant that lacks poly(ADP-ribose) glycohydrolase and have accumulated polyADP-ribosylated proteins in vivo, good extraction efficiency was obtained when the buffer contains 0.5% or more SDS. However it became also evident that polyADP-ribosylated proteins do not bind to the antibody column, presumably SDS did inhibit the binding. We had some success in eliminating the free SDS by adding the nonionic detergent in our preliminary experiments. Thus we are now trying to purify the polyADP-ribosylated proteins.
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[Journal Article] Inhibition of Crm1-p53 interaction and nuclear export of p53 by poly(ADP-ribosyl)ation2007
Author(s)
Kanai, M., Hanashiro, K., Kim, Song-Hee, Hanai, S., Boulares, A.H., Miwa, M. and Fukasawa, K.
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Journal Title
Nat.Cell Biol. 9
Pages: 1175-1183
Description
「研究成果報告書概要(和文)」より
Peer Reviewed
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[Journal Article] Inhibition of Crm1-p53 interaction and nuclear export of p53 by poly(ADP-ribosyl)ation.2007
Author(s)
Kanai, M., Hanashiro, K., Kim, Song-Hee, Hanai, S., Boulares, A. H., Miwa, M., Fukasawa, K.
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Journal Title
Nat. Cell Biol.(peer reviewed) 9
Pages: 1175-1183
Description
「研究成果報告書概要(欧文)」より
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