Base excision repair of the N-(2-deoxy-d-erythro-pentofuranosyl)-urea lesion by the hNEIL1 glycosylase

Rachana Tomar, Irina G. Minko, Pankaj Sharma, Andrew H. Kellum, Li Lei, Joel M. Harp, T. M. Iverson, R. Stephen Lloyd, Martin Egli, Michael P. Stone

Research output: Contribution to journalArticlepeer-review

6 Scopus citations

Abstract

The N-(2-deoxy-D-erythro-pentofuranosyl)-urea DNA lesion forms following hydrolytic fragmentation of cis-5R,6S-And trans-5R,6R-dihydroxy-5,6-dihydrothymidine (thymine glycol, Tg) or from oxidation of 7,8-dihydro-8-oxo-deoxyguanosine (8-oxodG) and subsequent hydrolysis. It interconverts between α and β deoxyribose anomers. Synthetic oligodeoxynucleotides containing this adduct are efficiently incised by unedited (K242) and edited (R242) forms of the hNEIL1 glycosylase. The structure of a complex between the active site unedited mutant Cδ100 P2G hNEIL1 (K242) glycosylase and double-stranded (ds) DNA containing a urea lesion reveals a pre-cleavage intermediate, in which the Gly2 N-Terminal amine forms a conjugate with the deoxyribose C1_ of the lesion, with the urea moiety remaining intact. This structure supports a proposed catalytic mechanism in which Glu3-mediated protonation of O4 facilitates attack at deoxyribose C1 . The deoxyribose is in the ring-opened configuration with the O4_ oxygen protonated. The electron density of Lys242 suggests the residue 242-in conformation associated with catalysis. This complex likely arises because the proton transfer steps involving Glu6 and Lys242 are hindered due to Glu6-mediated H-bonding with the Gly2 and the urea lesion. Consistent with crystallographic data, biochemical analyses show that the Cδ100 P2G hNEIL1 (K242) glycosylase exhibits a residual activity against urea-containing dsDNA.

Original languageEnglish (US)
Pages (from-to)3754-3769
Number of pages16
JournalNucleic acids research
Volume51
Issue number8
DOIs
StatePublished - May 8 2023

ASJC Scopus subject areas

  • Genetics

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