1996 Fiscal Year Final Research Report Summary
Probing DNA local structure by photoirradiation and application toward gene therapy.
| Project/Area Number |
07458143
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Bioorganic chemistry
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| Research Institution | KYOTO UNIVERSITY |
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Principal Investigator |
SUGIYAMA Hiroshi Kyoto University Engineering Associate Professor, 工学研究所, 助教授 (50183843)
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| Project Period (FY) |
1995 – 1996
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| Keywords | DNA cleavage / photo-electron-transfer / bromouracil / DNA structure / gene therapy / hydrogen abstraction |
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| Research Abstract |
In this research we focused on the fact that the hydrogen abstraction and electron transfer in DNA significantly depend on the local structures and sequence of DNA and we tried to find out a method to probe DNA local structure by the site of hydrogen abstraction and efficiency of electron transfer along DNA helix.
(1) In order to investigate the nature of deoxyribose C1' and C2' H abstraction by uracilyl-5-yl radical in B form DNA,we have examined the photoreaction of hexanucleotides d (GCA ^<Br>UGC) _2 in which one of the C2' hydrogens of the deoxyribose moiety was stereospecifically deuterated. We found that the formation of C2' oxidation product occurs via a rate-limiting abstraction of C2'alpha H of the deoxyribose.
(2) Photoreaction of 5-halouracil contained in a various biologically important DNA local structures, such as A,Z,and loop was examined. We found that in B DNA uracilyl-5-yl radical competitively abstract C1' and C2'alpha H of deoxyribose. In A form DNA,only C1' oxidation product was obtained whereas in Z-form DNA unprecedented C2' hydroxylation was oberved.
(3) In order to investigate the photoreactivity of ^<Br>U-containing DNA,uniformly ^<Br>U-substituted DNA was prepared by enzymatic primer extension method. The ^<Br>U-containing duplex DNA was irradiated at 302nm in the presence of iPrOH.It was found that the efficient cleavages occur at ^<Br>U residues located in 5'-A ^<Br>U^<Br>U-3' sites.
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