Detection of small structural changes of DNA under physiological conditions by a combination of site-selective deuteration and Raman difference spectroscopy
| Project/Area Number |
13672248
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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 |
Physical pharmacy
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| Research Institution | Tohoku University |
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Principal Investigator |
TOYAMA Akira Graduate School of Pharmaceutical Sciences, Assistant, 大学院・薬学研究科, 助手 (60217560)
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| Project Period (FY) |
2001 – 2002
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| Project Status |
Completed (Fiscal Year 2002)
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| Budget Amount *help |
¥3,500,000 (Direct Cost: ¥3,500,000)
Fiscal Year 2002: ¥700,000 (Direct Cost: ¥700,000)
Fiscal Year 2001: ¥2,800,000 (Direct Cost: ¥2,800,000)
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| Keywords | site-selective deuterated DNA / resonance Raman spectroscopy / nucleic acid structure / nucleic acid-drug interaction |
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| Research Abstract |
Small structural changes of selected guanine residues in DNA have been detected by a combination of site-selective isotope labeling and ultraviolet resonance Raman (UVRR) spectroscopy, I.e. isotope-edited Raman spectroscopy. UVRR spectra were measured for [8-^2H]guaninelabeled and unlabeled oligonucleotides excited at 251nm light. In the difference spectrum between the unlabeled and labeled DNA, Raman signals arise from the residue at the labeled position and the other components were canceled out. Thus, it becomes possible to extract the Raman signals of the selected guanine in olinonucleotides.
To obtain basic data for applying isotope-edited Raman spectroscopy to guanine residues, vibrational modes of UVRR bands of the C8-deuterated guanine ring were studied by examining the wavenumber shifts upon seven isotopic substitutions (2-^<13>C, 2-^<15>N, 6-^<18>O, 7-^<15>N, 8-^<13>C, 9-^<15>N and 1'-^<13>C). The hydrogen bond sensitivities of the Raman bands were also investigated by compari
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ng the Raman spectra recorded in several solvents of different hydrogen bonding properties. Some of the Raman bands have been found to be hydrogen bonding markers at specific donor or acceptor sites on the guanine ring, A negative peak around 1525 cm^<-1> a strong positive/negative peak pair around 1485/1465 cm^<-1>, and a weak positive/negative peak pair around 1025/1040cm^<-1> serve as markers of hydrogen bonding at N7, C6=O, and N1-H/C2-NH_2, respectively.
The applicability of isotopeedited Raman spectroscopy has been tested by using a 22-mer oligonucleotide duplex. UVRR spectra of the labeled and unlabeled oligomers were measured in the absence and presence of cAMP receptor protein (CRP),Which is Known to hydrogen-bonded to guanine residues of duplex DNA. The isotope-edited spectra showed that CRP is hydrogen-bonded strongly to C6=O of guanine residues but not N7 atom. Isotope-edited Raman spectroscopy can provide structural information of a selected guanine residue for DNA-drug systems under physiological conditions. Less
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Report
(3 results)
Research Products
(7 results)
