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

Jayati Roy Choudhury, Lu Rao, Ulrich Bierbach

Research output: Contribution to journalArticle

24 Citations (Scopus)

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(am 2)LCl](NO 3) 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 k obs = 1.4 ± 0.37 9 10 -4 s -1 (t 1/2 = 83 ± 22 min). Replacement of the thiourea group in ACRAMTU with an amidine group (compound 2) accelerates the rate by fourfold (k obs = 5.7 ± 0.58 9 10 -4 s -1, t 1/2 = 21 ± 2 min), and introduction of a propane-1,3- diamine nonleaving group results in a 1.5-fold enhancement in reactivity (compound 3, k obs = 2.1 ± 0.40 9 10 -4 s -1, t 1/2 = 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 2011
Externally publishedYes

Fingerprint

Intercalating Agents
Enzyme inhibition
Acridines
Assays
ethylenediamine
Diamines
DNA
Enzymes
Amidines
Thiourea
Propane
Endonucleases
Bioactivity
Platinum
Electrophoresis
Base Pairing
Nucleic Acids
Polyacrylamide Gel Electrophoresis
Rate constants
Pharmacology

Keywords

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

ASJC Scopus subject areas

  • Biochemistry
  • Inorganic Chemistry

Cite this

Rates of intercalator-driven platination of DNA determined by a restriction enzyme cleavage inhibition assay. / Choudhury, Jayati Roy; Rao, Lu; Bierbach, Ulrich.

In: Journal of Biological Inorganic Chemistry, Vol. 16, No. 3, 03.2011, p. 373-380.

Research output: Contribution to journalArticle

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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(am 2)LCl](NO 3) 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 k obs = 1.4 ± 0.37 9 10 -4 s -1 (t 1/2 = 83 ± 22 min). Replacement of the thiourea group in ACRAMTU with an amidine group (compound 2) accelerates the rate by fourfold (k obs = 5.7 ± 0.58 9 10 -4 s -1, t 1/2 = 21 ± 2 min), and introduction of a propane-1,3- diamine nonleaving group results in a 1.5-fold enhancement in reactivity (compound 3, k obs = 2.1 ± 0.40 9 10 -4 s -1, t 1/2 = 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.",
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N2 - 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(am 2)LCl](NO 3) 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 k obs = 1.4 ± 0.37 9 10 -4 s -1 (t 1/2 = 83 ± 22 min). Replacement of the thiourea group in ACRAMTU with an amidine group (compound 2) accelerates the rate by fourfold (k obs = 5.7 ± 0.58 9 10 -4 s -1, t 1/2 = 21 ± 2 min), and introduction of a propane-1,3- diamine nonleaving group results in a 1.5-fold enhancement in reactivity (compound 3, k obs = 2.1 ± 0.40 9 10 -4 s -1, t 1/2 = 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.

AB - 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(am 2)LCl](NO 3) 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 k obs = 1.4 ± 0.37 9 10 -4 s -1 (t 1/2 = 83 ± 22 min). Replacement of the thiourea group in ACRAMTU with an amidine group (compound 2) accelerates the rate by fourfold (k obs = 5.7 ± 0.58 9 10 -4 s -1, t 1/2 = 21 ± 2 min), and introduction of a propane-1,3- diamine nonleaving group results in a 1.5-fold enhancement in reactivity (compound 3, k obs = 2.1 ± 0.40 9 10 -4 s -1, t 1/2 = 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.

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