Chemical Modification of Intracellular Cytosine Deaminase from Chromobacterium violaceum YK 391

• Published in: Biotechnology and bioprocess engineering 2005, Vol.10 (3), p.180-185
• Main Authors: Kim, Jung, Kim, Tae-Hyun, Yu, Tae-Shick
• Format: Article
• Language: Korean
• ISSN: 1226-8372; 1976-3816

Cytosine deaminase (cytosine aminohydrolase, EC 3.5.4.1) stoichiometrically catalyzes the hydrolytic deamination of cytosine and 5-fluorocytosine to uracil and 5-fluorouracil, respectively. Amino acid residues located in or near the active sites of the intracellular cytosine deaminase from chromobacterium violaceum YK 391 were identified by chemical modification studies. The enzymic activity was completely inhibited by chemical modifiers, such as 1mM NBS, chloramine-T, $\rho-CMB,\;\rho-HMB$ and iodine, and was strongly inhibited by 1mM PMSF and pyridoxal 5'-phosphate. This chemical deactivation of the enzymic activity was reversed by a high concentration of cytosine. Furthermore, the deactivation of the enzymic activity by $\rho-CMB$ was also reversed by 1mM cysteine-HCI, DTT and 2-mercaptoethanol. These results suggested that cysteine, tryptophan and methionine residues might be located in or near the active sites of the enzyme, while serine and lysine were indirectly involved in the enzymic activity. The intracellular cytosine deaminase from C violaceum YK 391 was assumed to be a thiol enzyme.