| Fourier-transform infrared study of the photoactivation process of Xenopus (6-4) photolyase. | |
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MedLine Citation:
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PMID: 22747528 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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Photolyases (PHRs) are blue light-activated DNA repair enzymes that maintain genetic integrity by reverting UV-induced photoproducts into normal bases. The flavin adenine dinucleotide (FAD) chromophore of PHRs has four different redox states: oxidized (FAD(ox)), anion radical (FAD(•-)), neutral radical (FADH(•)), and fully reduced (FADH(-)). We combined difference Fourier-transform infrared (FTIR) spectroscopy with UV-visible spectroscopy to study the detailed photoactivation process of Xenopus (6-4) PHR. Two photons produce the enzymatically active, fully reduced PHR from oxidized FAD: FAD(ox) is converted to semiquinone via light-induced one-electron and one-proton transfers and then to FADH(-) by light-induced one-electron transfer. We successfully trapped FAD(•-) at 200 K, where electron transfer occurs but proton transfer does not. UV-visible spectroscopy following 450 nm illumination of FAD(ox) at 277 K defined the FADH(•)/FADH(-) mixture and allowed calculation of difference FTIR spectra among the four redox states. The absence of a characteristic C=O stretching vibration indicated that the proton donor is not a protonated carboxylic acid. Structural changes in Trp and Tyr are suggested by UV-visible and FTIR analysis of FAD(•-) at 200 K. Spectral analysis of amide I vibrations revealed structural perturbation of the protein's β-sheet during initial electron transfer (FAD(•-) formation), a transient increase in α-helicity during proton transfer (FADH(•) formation), and reversion to the initial amide I signal following subsequent electron transfer (FADH(-) formation). Consequently, in (6-4) PHR, unlike cryptochrome-DASH, formation of enzymatically active FADH(-) did not perturb α-helicity. Protein structural changes in the photoactivation of (6-4) PHR are discussed on the basis of these FTIR observations. |
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Authors:
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Daichi Yamada; Yu Zhang; Tatsuya Iwata; Kenichi Hitomi; Elizabeth D Getzoff; Hideki Kandori |
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Publication Detail:
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Type: Journal Article; Research Support, N.I.H., Extramural; Research Support, Non-U.S. Gov't Date: 2012-07-13 |
Journal Detail:
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Title: Biochemistry Volume: 51 ISSN: 1520-4995 ISO Abbreviation: Biochemistry Publication Date: 2012 Jul |
Date Detail:
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Created Date: 2012-07-24 Completed Date: 2012-10-01 Revised Date: 2012-11-01 |
Medline Journal Info:
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Nlm Unique ID: 0370623 Medline TA: Biochemistry Country: United States |
Other Details:
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Languages: eng Pagination: 5774-83 Citation Subset: IM |
Affiliation:
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Department of Frontier Materials, Nagoya Institute of Technology, Nagoya 466-8555, Japan. |
Export Citation:
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APA/MLA Format Download EndNote Download BibTex |
| MeSH Terms | |
Descriptor/Qualifier:
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Animals Deoxyribodipyrimidine Photo-Lyase / chemistry, metabolism* Flavin-Adenine Dinucleotide / chemistry, metabolism* Light Oxidation-Reduction Protein Structure, Secondary Spectrophotometry, Ultraviolet Spectroscopy, Fourier Transform Infrared / methods Xenopus / metabolism* Xenopus Proteins / chemistry, metabolism* |
| Grant Support | |
ID/Acronym/Agency:
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GM37684/GM/NIGMS NIH HHS; R01 GM037684/GM/NIGMS NIH HHS |
| Chemical | |
Reg. No./Substance:
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0/Xenopus Proteins; 146-14-5/Flavin-Adenine Dinucleotide; EC 4.1.99.3/Deoxyribodipyrimidine Photo-Lyase |
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