Application of chromatographic retention data in an investigation of a quantitative structure-nucleotide incorporation rate relationship

K. Valkó, T. Cserháti, I. Fellegvári, J. Sági, A. Szemzö

Research output: Contribution to journalArticle

12 Citations (Scopus)

Abstract

A series of 5-alkyl-, 5-alkenyl- and 5-alkynyl-substituted deoxyuridines and their triphosphate derivatives were synthesized and studied in DNA polymerase reactions. The initial rate of incorporation of the derivatives catalysed by Klenow fragment DNA polymerase enzyme (E. coli) was measured. Calf thymus DNA and synthetic poly (dA-dT) served as templates. The rate values were expressed as a percentage relative to the incorporation rate of natural substrate dTTP. The high-performance liquid chromatographic (HPLC) retention behaviours of the nucleoside derivatives were investigated on silica and reversed-phase stationary phases using various mixtures of ethyl acetate-methanol and methanol-water, as respectively, mobile phases. According to the results of principal component analysis, the HPLC retention data describe the hydrophobic properties of the compounds. The inclusion complex stability constants of the derivatives with cyclodextrins determined by reversed-phase thin-layer chromatography served as a measure of the steric properties of the subtituents. The electronic properties of the 5-substituents were characterized by the Swain-Lupton inductive and resonance parameters. The results of the stepwise linear regression analysis of the nucleotide incorporation rate data and the above-mentioned physico-chemical data revealed the importance of the electronic, steric and hydrophobic properties of the substituents in the DNA polymerase reactions. The importance of the steric parameter was more significant when the poly (dA-dT) template was used instead of the random base sequence template (calf thymus DNA).

Original languageEnglish
Pages (from-to)35-44
Number of pages10
JournalJournal of Chromatography A
Volume506
Issue numberC
DOIs
Publication statusPublished - May 11 1990

ASJC Scopus subject areas

  • Analytical Chemistry
  • Biochemistry
  • Organic Chemistry

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