New modified sarcosine oxidase, useful as diagnostic reagent for measuring creatinine or creatine, has reduced activity on N-ethylglycine, also nucleic acid that encodes it

摘要

Modified sarcosine oxidase (MSO) (I) which hydrolyzes 1 mole sarcosine to 1 mole each of glycine and formaldehyde; has reactivity for N-ethylglycine 70% or less of that of the wild-type enzyme; is active at pH 6.5-11, with optimum 8.0; has optimum temperature 55 [deg]C and is stable at 55 [deg]C or lower; and has apparent molecular weight 43 kD by sodium dodecylsulfate polyacrylamide gel electrophoresis, is new. Modified sarcosine oxidase (MSO) (I) comprises a 387 amino acid sequence (SEQ ID NO.1) fully defined in the specification, and which: (a) hydrolyzes 1 mole sarcosine to 1 mole each of glycine and formaldehyde; (b) has reactivity for N-ethylglycine 70% or less of that of the wild-type enzyme; (c) is active at pH 6.5-11, with optimum 8.0; (d) has optimum temperature 55 [deg]C and is stable at 55 [deg]C or lower; and (e) has apparent molecular weight 43 kD by sodium dodecylsulfate polyacrylamide gel electrophoresis; or its active derivatives with: one or more amino acid deletions, substitutions or additions; or 80% or more homology, provided reactivity for N-ethylglycine is lower than that of the wild-type enzyme. Independent claims are also included for: (1) gene (II) that encodes (I); (2) recombinant DNA (III) that comprises (II) incorporated into vector DNA; (3) transformants and transductants that include (III); (4) preparing (I); and (5) reagent for measuring creatinine or creatine that contains (I).