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Lesion search and recognition by thymine DNA glycosylase revealed by single molecule imaging

Please always quote using this URN: urn:nbn:de:bvb:20-opus-148795
  • The ability of DNA glycosylases to rapidly and efficiently detect lesions among a vast excess of nondamaged DNA bases is vitally important in base excision repair (BER). Here, we use singlemolecule imaging by atomic force microscopy (AFM) supported by a 2-aminopurine fluorescence base flipping assay to study damage search by human thymine DNA glycosylase (hTDG), which initiates BER of mutagenic and cytotoxic G:T and G:U mispairs in DNA. Our data reveal an equilibrium between two conformational states of hTDG-DNA complexes, assigned as searchThe ability of DNA glycosylases to rapidly and efficiently detect lesions among a vast excess of nondamaged DNA bases is vitally important in base excision repair (BER). Here, we use singlemolecule imaging by atomic force microscopy (AFM) supported by a 2-aminopurine fluorescence base flipping assay to study damage search by human thymine DNA glycosylase (hTDG), which initiates BER of mutagenic and cytotoxic G:T and G:U mispairs in DNA. Our data reveal an equilibrium between two conformational states of hTDG-DNA complexes, assigned as search complex (SC) and interrogation complex (IC), both at target lesions and undamaged DNA sites. Notably, for both hTDG and a second glycosylase, hOGG1, which recognizes structurally different 8-oxoguanine lesions, the conformation of the DNA in the SC mirrors innate structural properties of their respective target sites. In the IC, the DNA is sharply bent, as seen in crystal structures of hTDG lesion recognition complexes, which likely supports the base flipping required for lesion identification. Our results support a potentially general concept of sculpting of glycosylases to their targets, allowing them to exploit the energetic cost of DNA bending for initial lesion sensing, coupled with continuous (extrahelical) base interrogation during lesion search by DNA glycosylases.show moreshow less

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Metadaten
Author: Claudia N. Buechner, Atanu Maiti, Alexander C. Drohat, Ingrid Tessmer
URN:urn:nbn:de:bvb:20-opus-148795
Document Type:Journal article
Faculties:Fakultät für Biologie / Rudolf-Virchow-Zentrum
Language:English
Parent Title (English):Nucleic Acids Research
Year of Completion:2015
Volume:43
Issue:5
Pagenumber:2716-2729
Source:Nucleic Acids Research, 2015, Vol. 43, No. 5. DOI: 10.1093/nar/gkv139
DOI:https://doi.org/10.1093/nar/gkv139
Dewey Decimal Classification:6 Technik, Medizin, angewandte Wissenschaften / 61 Medizin und Gesundheit / 610 Medizin und Gesundheit
Tag:Escherichia coli AlkA; base excision repair; damaged DNA; intrahelical lesion; mismatch recognition; structural basis; substrate recognition; undamaged DNA; uracil binding
Release Date:2018/11/21
Licence (German):License LogoCC BY: Creative-Commons-Lizenz: Namensnennung 4.0 International