Inhibition by aphidicolin and dideoxythymidine triphosphate of a multienzyme complex of DNA synthesis from human cells.
A multienzyme complex consisting of DNA polymerase and several DNA precursor-synthesizing enzymes was solubilized by gentle lysis of cultured human cells. This complex channelled the distal precursor [3H]dTMP into DNA. The patterns of inhibition of the complex by aphidicolin and dideoxythymidine triphosphate (ddTTP) suggested that the complex contained the replicative DNA polymerase, polymerase alpha. Inhibition by ddTTP was competitive with dTTP. This was exploited to estimate the effective concentration of [3H]dTTP at the site of DNA synthesis during channelling of [3H]dTMP into DNA. The estimated concentration (about 50 microM) was so high as to suggest that the solubilized complex was able to functionally compartmentalize DNA precursors.
D016590:Aphidicolin; D004261:DNA Replication / Q000187:drug effects*; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism; D054306:Dideoxynucleotides; D004224:Diterpenes / Q000494:pharmacology*; D006801:Humans; D009097:Multienzyme Complexes / Q000378:metabolism; D013942:Thymine Nucleotides / Q000378:metabolism / Q000494:pharmacology*
D054306:Dideoxynucleotides; D004224:Diterpenes; D009097:Multienzyme Complexes; D013942:Thymine Nucleotides; D016590:Aphidicolin; D004259:DNA-Directed DNA Polymerase; C017216:2',3'-dideoxythymidine triphosphate; C024157:thymidine 5'-triphosphate