Benzo[c]phenanthrene adducts and nogalamycin inhibit DNA transesterification by vaccinia topoisomerase Journal Article
| Authors: | Yakovleva, L.; Handy, C. J.; Sayer, J. M.; Pirrung, M.; Jerina, D. M.; Shuman, S. |
| Article Title: | Benzo[c]phenanthrene adducts and nogalamycin inhibit DNA transesterification by vaccinia topoisomerase |
| Abstract: | Vaccinia DNA topoisomerase forms a covalent DNA-(3′ -phosphotyrosyl)-enzyme intermediate at a specific target site 5′-C +5C+4C+3T+2T+1p ↓ N-1 in duplex DNA. Here we study the effects of position-specific DNA intercalators on the rate and extent of single-turnover DNA transesterification. Chiral C-1 R and S trans-opened 3,4-diol 1,2-epoxide adducts of benzo[c]phenanthrene (BcPh) were introduced at single N 2-deoxyguanosine and N6-deoxyadenosine positions within the 3′-G+5G+4G+3A+2A +1-T+1A-2 sequence of the nonscissile DNA strand. Transesterification was unaffected by BcPh intercalation between the +6 and +5 base pairs, slowed 4-fold by intercalation between the +5 and +4 base pairs, and virtually abolished by BcPh intercalation between the +4 and +3 base pairs and the +3 and +2 base pairs. Intercalation between the +2 and +1 base pairs by the +2R BcPh dA adduct abolished transesterification, whereas the overlapping +1S BcPh dA adduct slowed the rate of transesterification by a factor of 2700, with little effect upon the extent of the reaction. Intercalation at the scissile phosphodiester (between the +1 and -1 base pairs) slowed transesterification by a factor of 450. BcPh intercalation between the -1 and -2 base pairs slowed cleavage by two orders of magnitude, but intercalation between the -2 and -3 base pairs had little effect. The anthracycline drug nogalamycin, a noncovalent intercalator with preference for 5′-TG dinucleotides, inhibited the single-turnover DNA cleavage reaction of vaccinia topoisomerase with an IC50 of 0.7 μM. Nogalamycin was most effective when the drug was preincubated with DNA and when the cleavage target site was 5 ′-CCCTT ↓ G instead of 5′-CCCTT ↓ A. These findings demarcate upstream and downstream boundaries of the functional interface of vaccinia topoisomerase with its DNA target site. |
| Keywords: | unclassified drug; nonhuman; gene targeting; dna; double stranded dna; molecular sequence data; molecular recognition; vaccinia virus; base sequence; base pairing; dna adduct; dna adducts; models, molecular; esterification; nucleic acid conformation; protein folding; dna topoisomerase inhibitor; anthracycline; enzyme specificity; biochemistry; hydrolysis; enzymes; dna synthesis inhibition; enzyme mechanism; transesterification; dna cleavage; dna topoisomerases, type i; vaccinia; chirality; drug products; covalent bond; adducts; target sites; aromatic compounds; intercalation compounds; deoxyadenosine; intercalating agents; phenanthrene derivative; benzo[c]phenanthridine derivative; phenanthrenes; priority journal; article; benzo[c]phenanthrene; nogalamycin; staphylococcus phage 3a |
| Journal Title: | Journal of Biological Chemistry |
| Volume: | 279 |
| Issue: | 22 |
| ISSN: | 0021-9258 |
| Publisher: | American Society for Biochemistry and Molecular Biology |
| Date Published: | 2004-05-28 |
| Start Page: | 23335 |
| End Page: | 23342 |
| Language: | English |
| DOI: | 10.1074/jbc.M401203200 |
| PROVIDER: | scopus |
| PUBMED: | 15044474 |
| DOI/URL: | |
| Notes: | J. Biol. Chem. -- Cited By (since 1996):16 -- Export Date: 16 June 2014 -- CODEN: JBCHA -- Source: Scopus |
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