Method for producing nucleoside-5′-phosphate ester
AssigneeAjinomoto Co., Inc.
InventorsYasuhiro Mihara, Takashi Utagawa, Hideaki Yamada, Yasuhisa Asano
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Abstract
A method for producing nucleoside-5′-phosphate esters inexpensively and in high yields by phosphorylating a nucleoside with a phospahte group donor using an acid phosphatase having an increased affinity for the nucleoside and/or an increased temperature stability at a pH of pH 3.0 to 5.5, to produce a nucleoside-5′-phosphate ester. Mutant acid phosphatases having increased affinity for nucleosides and/or an enhanced temperature stability are also provided.
Claims3
A mutant acid phosphatase wherein the wild-type acid phosphatase comprising an amino acid sequence selected from the group consisting of SEQ ID Nos:
- 4, 8, 24, 26, and 28, and said acid phosphatase has a mutation which decreases the Km value for a nucleoside and/or increases the temperature stability as compared to said wild-type acid phosphatase;
- wherein said nucleoside is selected from the group consisting of inosine, guanosine, adenosine, xanthosine, purine riboside, 6-methoxypurine riboside, 2,6-diaminopurine riboside, 6-fluoropurine riboside, 6-thiopurine riboside, 2-amino-6-thiopurine riboside, mercaptoguanosine uridine, cytidine, 5-aminouridine, 5-hydroxyuridine, 5-bromouridine, and 6-azauridine;
- and wherein the Km value for the nucleoside is determined by measuring the transphosphorylation activity of the acid phosphatase at 30° C. and pH4.0 in the presence of 100 μmol/ml of sodium pyrophosphate.
Dependencies are inferred from the claim text.