Specific selection of deoxycytidine kinase mutants with tritiated deoxyadenosine

Teh sheng Chan, J. Arly Nelson

Research output: Contribution to journalArticle

Abstract

We have shown previously that a low concentration of tritiated deoxyadenosine, i.e., 1 μCi/ml, selectively kills wild-type S49 murine lymphoma cells. Mutant cells resistant to [3H]deoxyadenosine lacked adenosine kinase completely but retained a significant level of deoxyadenosine phosphorylating activity. To study further the specificity of [3H]deoxyadenosine selection, lymphoma cell clones resistant to 15 μCi/ml [3H]deoxyadenosine have been derived. The resistant line, S49-dA15, is also resistant to high levels of nonradioactive deoxyadenosine and to deoxyguanosine but remains sensitive to thymidine. The thymidine inhibition of the growth of the mutant, in contrast to that of the wild-type cells, cannot be prevented by deoxycytidine. The mutant line lacks deoxycytidine kinase that also phosphorylates deoxyadenosine. In addition, the mutant cells excrete a large amount of deoxycytidine into culture medium, consistent with a failure of salvage of the nucleoside in the absence of an appropriate kinase, i.e., deoxycytidine kinase. In contrast, a deoxycytidine kinase-deficient cell line that was selected with arabinosylcytosine does not excrete deoxycytidine and contains high deoxycytidine deaminase activity. [3H]Deoxyadenosine can be used as a selective agent for specific selection of deoxycytidine kinase-negative mutants.

Original languageEnglish (US)
Pages (from-to)327-340
Number of pages14
JournalBiochemical Genetics
Volume33
Issue number9-10
DOIs
StatePublished - Oct 1995

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Keywords

  • adenosine kinase
  • deoxycytidine kinase
  • nucleoside analogue
  • tritium suicide

ASJC Scopus subject areas

  • Genetics
  • Biochemistry

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