Perturbation of ATP-induced tetramerization of human cytosolic thymidine kinase by substitution of serine-13 with aspartic acid at the mitotic phosphorylation site |
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Authors: | Li Chia-Lung Lu Ching-Yi Ke Po-Yuan Chang Zee-Fen |
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Affiliation: | Institute of Biochemistry and Molecular Biology, College of Medicine, National Taiwan University, No 1 Jen Ai Road First Section, Taipei, Taiwan ROC. |
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Abstract: | Human cytosolic thymidine kinase (TK1) is tightly regulated in the cell cycle by multiple mechanisms. Our laboratory has previously shown that in mitotic-arrested cells human TK1 is phosphorylated at serine-13, accompanied by a decrease in catalytic efficiency. In this study we investigated whether serine-13 phosphorylation regulated TK1 activity and found that substitution of serine-13 with aspartic acid (S13D), which mimics phosphorylation, not only diminished the ATP-activating effect on the enzyme, but also decreased its thymidine substrate affinity. Our experimental results further showed that the S13D mutation perturbed ATP-induced tetramerization of TK1. Given that the dimeric form of TK1 is less active than the tetrameric, we propose that mitotic phosphorylation of serine-13 is of physiological importance, in that it may counteract ATP-dependent activation of TK1 by affecting its quaternary structure, thus attenuating its enzymatic function at the G2/M phase. |
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Keywords: | Phosphorylation Thymidine kinase Cell cycle Kinetics Oligomerization |
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