Effect of C-terminal of human cytosolic thymidine kinase (TK1) on in vitro stability and enzymatic properties.
Thymidine kinase (TK1) is a key enzyme in the salvage pathway of nucleotide metabolism and catalyzes the first rate-limiting step in the synthesis of dTTP, transfer of a gamma-phosphate group from a nucleoside triphosphate to the 5'-hydroxyl group of thymidine, thus forming dTMP. TK1 is cytosolic and its activity fluctuates during cell cycle coinciding with the DNA synthesis rate and disappears during mitosis. This fluctuation is important for providing a balanced supply of dTTP for DNA replication.The cell cycle specific activity of TK1 is regulated at the transcriptional level, but posttranslational mechanisms seem to play an important role for the level of functional TK1 protein as well. Thus, the C-terminal of TK1 is known to be essential for the specific degradation of the enzyme at the G2/M phase. In this work, we have studied the effect of deletion of the C-terminal 20, 40, and 44 amino acids of TK1 on in vitro stability, oligomerization, and enzyme kinetics. We found that deletion of the C-terminal fold markedly increased the stability as well as the catalytic activity.
Zhu
C
C
Department of Life Sciences and Chemistry, Roskilde University, Roskilde, Denmark.
Harlow
L S
LS
Berenstein
D
D
Munch-Petersen
S
S
Munch-Petersen
B
B
D002384:Catalysis; D002453:Cell Cycle; D002455:Cell Division; D003600:Cytosol / Q000201:enzymology* / Q000378:metabolism; D004247:DNA / Q000737:chemistry; D004261:DNA Replication; D016195:G2 Phase; D017353:Gene Deletion; D006801:Humans; D066298:In Vitro Techniques; D008938:Mitosis; D017434:Protein Structure, Tertiary; D013937:Thymidine Kinase / Q000096:biosynthesis*; D014158:Transcription, Genetic
D004247:DNA; D013937:Thymidine Kinase; C092717:thymidine kinase 1