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Esstntial amino acids involved in the catalytic role of the extracellular cytosine deaminase from Chromobacterium violaceum YK 391 were determined by chemical modification studies. The enzyme activity required the reduced form of Fe (Ⅱ) ion, since the enzyme was inhibited by ο-phenanthroline. The enzyme activity was completely inhibited by the chemical modifiers, such as p-chloromercuribenzoate (p-CMB), phydroxymercuribenzoate, and chloramine-T at 1 mM each. The enzyme activity was also markedly inhibited by pyridoxal-5´-phosphate, diethyl pyrocarbonate, and phenylmethylsulfonyl fluroride at 1 mM each. The inactivation of the enzyme activity with p-CMB was reversed by a high concentration of cytosine. Furthermore, the inactivation of the enzyme activity with p-CMB was also reactivated by 1 mM dithiothreitol, 1 mM 2-mercaptoethanol, 1 mM cysteine-HCl, 10% ethyl alcohol, and 10% methyl alcohol. These results suggested that cysteine and methionine residues might be located in or near the active site of the enzyme, while lysine, histidine, and serine residues might be indirectly involved in the enzyme activity.
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