Benzo[a]pyrene-dG adduct interference illuminates the interface of vaccinia topoisomerase with the DNA minor groove Journal Article
| Authors: | Tian, L.; Sayer, J. M.; Kroth, H.; Kalena, G.; Jerina, D. M.; Shuman, S. |
| Article Title: | Benzo[a]pyrene-dG adduct interference illuminates the interface of vaccinia topoisomerase with the DNA minor groove |
| Abstract: | Vaccinia DNA topoisomerase forms a covalent DNA-(3′-phosphotyrosyl)-enzyme intermediate at a pentapyrimidine target site 5′-C+5C+4C+3T+2 T+1p ↓ in duplex DNA. The enzyme engages the target site within a C-shaped protein clamp. Here we mapped the interface of topoisomerase with the DNA minor groove by introducing chiral C-10 R and S 7,8-diol 9,10-epoxide adducts of benzo[a]pyrene (BP) at single N2-deoxyguanosine (dG) positions within the nonscissile DNA strand. These trans opened BPdG adducts fit into the minor groove without perturbing helix conformation or base pairing, and the R and S diastereomers are oriented in opposite directions within the minor groove. We measured the effects of the BPdG adducts on the rate and extent of single-turnover DNA transesterification. We observed a sharp margin of interference effects, whereby +5 and -2 BPdG modifications were well tolerated but +4, +3, and -1 BPdG adducts were severely deleterious. Stereoselective effects at the -1 nucleoside (the R isomer interfered, whereas the S isomer did not) delineated at high resolution the downstream border of the minor groove interface. BPdG inhibition of transesterification is likely caused by steric exclusion of constituents of the topoisomerase from the minor groove. We also applied the BPdG interference method to probe the interactions of exonuclease III with the minor groove. DNAs containing these BPdG adducts were protected from digestion by exonuclease III, which was consistently arrested at positions 2-4 nucleotides prior to the BP-modified guanosine. |
| Keywords: | nonhuman; molecular genetics; metabolism; protein dna binding; protein binding; enzymology; time; time factors; chemistry; dna; double stranded dna; molecular sequence data; kinetics; dna viruses; nucleotide sequence; vaccinia virus; base sequence; binding site; benzo[a]pyrene; deoxyguanosine; base pairing; dna adduct; benzo(a)pyrene; dna adducts; models, molecular; binding sites; esterification; chemical structure; high performance liquid chromatography; chromatography, high pressure liquid; conformation; nucleic acid conformation; models, chemical; enzymes; transesterification; stereochemistry; nucleoside; dna topoisomerase; dna topoisomerases, type i; exodeoxyribonuclease; exodeoxyribonucleases; vaccinia; conformations; diastereoisomer; chirality; chemical model; dna strand; isomers; exodeoxyribonuclease iii; priority journal; article; diastereomers; benzo(a)pyrene dna adduct; benzo(a)pyrene-dna adduct |
| Journal Title: | Journal of Biological Chemistry |
| Volume: | 278 |
| Issue: | 11 |
| ISSN: | 0021-9258 |
| Publisher: | American Society for Biochemistry and Molecular Biology |
| Date Published: | 2003-03-14 |
| Start Page: | 9905 |
| End Page: | 9911 |
| Language: | English |
| DOI: | 10.1074/jbc.M212468200 |
| PUBMED: | 12524450 |
| PROVIDER: | scopus |
| DOI/URL: | |
| Notes: | Export Date: 12 September 2014 -- Source: Scopus |
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