Teruo Arima

Alternative methods of thymidine phosphorylation in different organisms

Abstract Thymidine was found to be phosphorylated to thymidine 5′-monophosphate using ATP as phosphate donor in germinating potatoes and Tetrahymena pyriformis as well as in regenerating rat liver and tumor tissues. The properties of the phosphorylating enzymes from these sources were studied. The results obtained suggest that germinating potatoes and T. pyriformis may not contain thymidine kinase (ATP : thymidine 5′-monophosphotransferase, EC 2.7.1.21). In germinating potatoes and T. pyriformis , thymidine was phosphorylated by the combined actions of nucleoside phosphotransferase and the ATP hydrolyzing enzyme. The nucleoside phosphotransferase and ATP hydrolyzing enzyme from germinating potatoes were separated by DEAE-Sephadex A-50 column chromatography, but these enzymes from T. pyriformis could not be separated under the same conditions. When an extract of T. pyriformis was treated with trypsin, both the thymidine phosphorylating activities using ATP as phosphate donor and ATP hydrolyzing activities disappeared, while the nucleoside phosphotransferase activity remained. These results suggest that thymidine can be phosphorylated in two ways: one is by the action of thymidine kinase (in mammalian tissues and Escherichia coli ) and the other is by the combined actions of nucleoside phosphotransferase and the ATP hydrolyzing enzyme (in potatoes and T. pyriformis ).

Studies on a reciprocal relationship between nucleoside kinases and 5′-nucleotidase

Abstract The activities of thymidine kinase (EC 2.7.1.21), deoxycytidine kinase, uridine kinase (EC 2.7.1.48) and 5′-nucleotidase (EC 3.1.3.5) for dTMP, dCMP and UMP were investigated during liver regeneration. A reciprocal relationship was found between the activity of thymidine kinase and 5′-nucleotidase activity for dTMP during liver regeneration. This relationship was also found in various normal and tumor tissues. Similar relationships were also found between uridine kinase and 5′-nucleotidase for UMP and between deoxycytidine kinase and 5′-nucleotidase for dCMP during liver regeneration. Treatment of animals with actinomycin D inhibited the elevation of thymidine kinase activity during liver regeneration, but had no effect on that of uridine kinase. This treatment also inhibited the decreases of 5′-nucleotidase activity for dTMP and UMP during liver regeneration. The thymidine kinase and uridine kinase activities of regenerating liver were inhibited by addition of the microsomal fraction from normal liver. The microsomal fraction from regenerating liver was less inhibitory. The microsomal fraction contained various phosphatases, such as 5′-nucleotidase and ATPase, and it seems probable that the inhibotory effect of this fraction on the kinases may result from the actions of these phosphatases. It was found that the activities of the phosphatases in the microsomal fraction of regenerating liver were much lower than those of normal liver.

Studies on aminopeptidases in rat liver and plasma

Abstract It was found that aminopeptidases (EC 3.4.1.2) were easily solubilized from rat liver by bromelain (EC 3.4.4.24) treatment. The solubilized form of the enzymes was subject to TEAE-cellulose column chromatography resulting in separation of five types of aminopeptidases. Rat plasma also contained some aminopeptidases, which were separated by gel filtration on a Sephadex G-200 column or TEAE-cellulose column chromatography. These enzymes showed different substrate specificities toward l -leucine amide and l -leucyl-β-naphthylamide. It was found that these enzymes increased in the plasma after CCl 4 injection or ligation of the common bile duct.

Inhibitor of pyrimidine metabolism from tumor tissues

Abstract Inhibitors of normal rat liver 5′-nucleotidase and dUMP kinase in vitro were found in rapidly proliferating tissues, such as Yoshida sarcoma. Two inhibitors were separated from Yoshida sarcoma by zone electrophoresis, gel filtration on Sephadex G-200 and DEAE-cellulose column chromatography. One inhibited both 5′-nucleotidase and dUMP kinase, while the other inhibited only dUMP kinase. These inhibitors were not detectable in normal rat liver. They were induced in regenerating rat liver and present in rapidly proliferating tissues, such as Yoshida sarcoma and Ehrlich ascites tumor and rat marrow cells. These inhibitors were heat labile. One had a large molecular weight (500,000>) and the other a small molecular weight ( Ca . 50,000).