Cooperative inhibition of human thymidylate synthase by mixtures of active site binding and allosteric inhibitors

被引:34
|
作者
Lovelace, Leslie L.
Gibson, Lydia M.
Lebioda, Lukasz [1 ]
机构
[1] Univ S Carolina, Dept Chem & Biochem, Columbia, SC 29208 USA
[2] Univ S Carolina, Ctr Colon Canc Res, Columbia, SC 29208 USA
关键词
D O I
10.1021/bi061309j
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Thymidylate synthase (TS) is a target in the chemotherapy of colorectal cancer and some other neoplasms. It catalyzes the transfer of a methyl group from methylenetetrahydrofolate to dUMP to form dTMP. On the basis of structural considerations, we have introduced 1,3-propanediphosphonic acid (PDPA) as an allosteric inhibitor of human TS (hTS); it is proposed that PDPA acts by stabilizing an inactive conformer of loop 181-197. Kinetic studies showed that PDPA is a mixed (noncompetitive) inhibitor versus dUMP. In contrast, versus methylenetrahydrofolate at concentrations lower than 0.25 AM, PDPA is an uncompetitive inhibitor, while at PDPA concentrations higher than 1 mu M the inhibiton is noncompetive, as expected. At the concentrations corresponding to uncompetitive inhibition, PDPA shows positive cooperativity with an antifolate inhibitor, ZD9331, which binds to the active conformer. PDPA binding leads to the formation of hTS tetramers, but not higher oligomers. These data are consistent with a model in which hTS exists preferably as an asymmetric dimer with one subunit in the active conformation of loop 181-197 and the other in the inactive conformation.
引用
收藏
页码:2823 / 2830
页数:8
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