INCORPORATION OF 5-AZA-2'-DEOXYCYTIDINE-5'-TRIPHOSPHATE INTO DNA - INTERACTIONS WITH MAMMALIAN DNA POLYMERASE-ALPHA AND DNA METHYLASE

  • 1 January 1983
    • journal article
    • research article
    • Vol. 24  (1) , 109-114
Abstract
To understand further the molecular mode of action of 5-Aza-2''-deoxycytidine (5-AZA-dCyd), a potent antileukemic agent, 5-Aza-2''-deoxycytidine 5''-triphosphate (5-AZA-dCTP), was enzymatically prepared, and studies were performed with purified DNA polymerase .alpha. and DNA methylase from mammalian cells. DNA polymerase .alpha. catalyzed the incorporation of 5-AZA-dCTP into DNA. The apparent Km for 5-AZA-dCTP was estimated to be 3.0 .mu.M; the Km of dCTP was 2.0 .mu.M. The apparent Vmax of 5-AZA-dCTP was slightly lower than that for dCTP. 5-AZA-dCTP was a weak, competitive inhibitor (Ki 4.3 .mu.M) with respect to dCTP. Template studies with 5-AZA-dCTP showed that this nucleotide analog was incorporated into poly(dIC), but not into poly(dAT), suggesting that the incorporation follows the rules of Watson-Crick base pairing. Incorporation of 5-AZA-dCTP into hemimethylated DNA produced a significant inhibition of DNA methylase. 5-AZA-dCTP is a very good substrate for DNA polymerase .alpha., and its incorporation into DNA inhibits DNA methylation.