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Inhibition by aphidicolin and dideoxythymidine triphosphate of a multienzyme complex of DNA synthesis from human cells
Author(s) -
Wickremasinghe R.Gitendra,
Hoffbrand A.Victor
Publication year - 1983
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/0014-5793(83)80441-4
Subject(s) - aphidicolin , dna synthesis , dna , dna polymerase , microbiology and biotechnology , biology , dna clamp , lysis , biochemistry , dna polymerase ii , enzyme , thymidine , polymerase , chemistry , polymerase chain reaction , reverse transcriptase , gene
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 [ 3 H]dTMP into DNA. The patterns of inhibition of the complex by aphidicolin and dideoxythymidine triphopshate (ddTTP) suggested that the complex contained the replicative DNA polymerase, polymerase α. Inhibition by ddTTP was competitive with dTTP. This was exploited to estimate the effective concentration of [ 3 H]dTTP at the site of DNA synthesis during channelling of [ 3 H]dTMP into DNA. The estimated concentration (about 50 μM) was so high as to suggest that the solubilized complex was able to functionally compartmentalize DNA precursors.