Rates of intercalator-driven platination of DNA determined by a restriction enzyme cleavage inhibition assay

Jayati Roy Choudhury, Lu Rao, Ulrich Bierbach

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Abstract

A restriction enzyme cleavage inhibition assay was designed to determine the rates of DNA platination by four non-cross-linking platinum-acridine agents represented by the formula [Pt(am2)LCl](NO3)2, where am is a diamine nonleaving group and L is an acridine derived from the intercalator 1-[2-(acridin-9-ylamino)ethyl]-1,3-dimethylthiourea (ACRAMTU). The formation of monofunctional adducts in the target sequence 5′-CGA was studied in a 40-base-pair probe containing the EcoRI restriction site GAATTC. The time dependence of endonuclease inhibition was quantitatively analyzed by polyacrylamide gel electrophoresis. The formation of monoadducts is approximately 3 times faster with double-stranded DNA than with simple nucleic acid fragments. Compound 1 (am2 is ethane- 1,2-diamine, L is ACRAMTU) reacts with a first-order rate constant of kobs = 1.4 ± 0.37 9 10-4 s-1 (t1/2 = 83 ± 22 min). Replacement of the thiourea group in ACRAMTU with an amidine group (compound 2) accelerates the rate by fourfold (kobs = 5.7 ± 0.58 9 10 -4 s-1, t1/2 = 21 ± 2 min), and introduction of a propane-1,3- diamine nonleaving group results in a 1.5-fold enhancement in reactivity (compound 3, kobs = 2.1 ± 0.40 9 10-4 s-1, t1/2 = 55 ± 10 min) compared with the prototype. Derivative 4, containing a 4,9-disubstituted acridine threading intercalator, was the least reactive compound in the series (k obs = 1.1 ± 0.40 9 10-4 s-1, t 1/2 = 104 ± 38 min). The data suggest a correlation may exist between the binding rates and the biological activity of the compounds. Potential pharmacological advantages of rapid formation of cytotoxic monofunctional adducts over the common purine-purine cross-links are discussed.

Original languageEnglish (US)
Pages (from-to)373-380
Number of pages8
JournalJournal of Biological Inorganic Chemistry
Volume16
Issue number3
DOIs
StatePublished - Mar 1 2011
Externally publishedYes

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Keywords

  • DNA binding
  • Gel electrophoresis
  • Kinetics
  • Monofunctional adducts
  • Platinum-acridine agents

ASJC Scopus subject areas

  • Biochemistry
  • Inorganic Chemistry

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