期刊
CHEMBIOCHEM
卷 17, 期 23, 页码 2240-2249出版社
WILEY-V C H VERLAG GMBH
DOI: 10.1002/cbic.201600405
关键词
antitumor agents; fluorinated nucleotides; gemcitabine; inhibitors; molecular recognition
资金
- Canadian Institutes of Health Research
- Scotia Support Grant from the Nova Scotia Health Research Foundation
- Discovery Grant from the Natural Sciences and Engineering Research Council of Canada
- Craig's Cause Pancreatic Cancer Society
- CIBC Graduate Scholarship in Medical Research
- Terry Fox Research Institute
CTP synthase (CTPS) catalyzes the conversion of UTP to CTP and is a target for the development of antiviral, anticancer, antiprotozoal, and immunosuppressive agents. Exposure of cell lines to the antineoplastic cytidine analogue gemcitabine causes depletion of intracellular CTP levels, but the direct inhibition of CTPS by its metabolite gemcitabine-5'-triphosphate (dF-dCTP) has not been demonstrated. We show that dF-dCTP is a potent competitive inhibitor of Escherichia coli CTPS with respect to UTP [K-i=(3.0 +/- 0.1) mu m], and that its binding affinity exceeds that of CTP approximate to 75-fold. Site-directed mutagenesis studies indicated that Glu149 is an important binding determinant for both CTP and dF-dCTP. Comparison of the binding affinities of the 5'-triphosphates of 2'-fluoro-2'-deoxycytidine and 2'-fluoro-2'-deoxyarabinocytidine revealed that the 2'-F-arabino group contributes markedly to the strong binding of dF-dCTP. Geminal 2'-F substitution on UTP (dF-dUTP) did not result in an increase in binding affinity with CTPS. Remarkably, CTPS catalyzed the conversion of dF-dUTP into dF-dCTP, thus suggesting that dF-dCTP might be regenerated in vivo from its catabolite dF-dUTP.
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