A simple and sensitive ribonucleotide reductase assay

Ambrose Y. Jong, Kefei Yu, Bingsen Zhou, Tomǎs Frgala, Patrick Reynolds, Yun Yen

研究成果: 雜誌貢獻文章同行評審

17 引文 斯高帕斯(Scopus)


Ribonucleotide reductase (RR) is a key regulatory enzyme in the DNA synthesis pathway and is the target of the cancer chemotherapeutic agent hydroxyurea. The study of RR is significantly hindered by the tedious and labor-intensive nature of enzymatic assay. In this report, we present a novel RR assay in which detection of the deoxyribonucleotides produced by RR occurs via coupling to the DNA polymerase reaction, and is enhanced by using RNase to degrade endogenous RNA. Cell extracts from various cell lines were treated with RNase and then reacted with ATP and radioactive ribonucleotide diphosphate as the substrate. Incorporation of the radioactive substrate [14C]CDP into DNA was linear over 30 min and was linear with the amount of extract, which provided RR activity. The reaction was inhibited by hydroxyurea and required Mg2+ and ATP, suggesting that the assay is specific to RR activity. While RR activities determined by our method and by a conventional method were comparable, this novel method proved to be simpler faster, more sensitive and less expensive. In addition, assay of the RR activity for multiple samples can easily be performed simultaneously. It is superior to other RR assays in all aspects.

頁(從 - 到)62-68
期刊Journal of Biomedical Science
出版狀態已發佈 - 1998

ASJC Scopus subject areas

  • 內分泌學、糖尿病和代謝
  • 分子生物學
  • 臨床生物化學
  • 細胞生物學
  • 生物化學(醫學)
  • 藥學(醫學)


深入研究「A simple and sensitive ribonucleotide reductase assay」主題。共同形成了獨特的指紋。