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    • 7. 发明授权
    • Urinary kallikrein assay: specific substrates and assay method
    • 尿激肽释放酶测定:特异性底物和测定方法
    • US4327178A
    • 1982-04-27
    • US249645
    • 1981-03-31
    • James W. RyanAlfred Chung
    • James W. RyanAlfred Chung
    • C12Q1/37C12Q1/38C12Q1/36
    • C12Q1/37Y10S530/802
    • Substrates and method for the assay of urinary kallikrein are provided. The substrates have the general formula ##STR1## wherein R is H, acetyl, benzoyl, cyclopentylcarbonyl, succinyl, R.sub.1 --Ser.sub.1 or R.sub.1 --Phe--Ser where R.sub.1 is H, acetyl, benzoyl, cyclopentylcarbonyl or succinyl,X is H, tritium, 3-iodo or 4-iodo, andn is 0 or 1.Radioactive label may be incorporated in the anilide or benzylamide moiety. Hydrolysis catalyzed by urinary kallikrein yields labeled aniline or benzylamine as product.The assay method includes mixing the enzyme with substrate in a buffered solution, pH 7.5-10.5, the substrate being present preferably at a concentration substantially below K.sub.m. After incubating to allow the reaction to proceed, the reaction is terminated and the radioactive hydrolysis product is separated and counted.Also disclosed are compounds of the type ##STR2## wherein R, R.sub.1 and n are as defined above. X is 3--OH or 4--OH. Y.sub.1 is 3-iodo if X is 4--OH or 6-iodo if X is 3--OH. Y.sub.2 may be H or 5-iodo if Y.sub.1 is 3-iodo, or 4-iodo if Y.sub.1 is 6-iodo.
    • 提供尿激肽释放酶的底物和方法。 底物具有通式:其中R为H,乙酰基,苯甲酰基,环戊基羰基,琥珀酰基,R1-Ser1或R1-Phe-Ser,其中R1为H,乙酰基,苯甲酰基,环戊基羰基或琥珀酰基,X为H,氚, 3-碘或4-碘,n为0或1.放射性标记可以并入苯胺或苄基酰胺部分。 通过尿激肽释放酶催化水解产生标记的苯胺或苄胺作为产物。 测定方法包括将酶与底物在缓冲溶液(pH7.5-10.5)中混合,底物优选以基本上低于Km的浓度存在。 孵育后使反应进行,终止反应,分离并计数放射性水解产物。 还公开了类型“IMAGE”的化合物,其中R,R 1和n如上所定义。 X是3-OH或4-OH。 如果X为3-OH,X为4-羟基或6-碘,则Y1为3-碘。 如果Y1是3-碘,Y2可以是H或5-碘,如果Y1是6-碘,Y2可以是4-碘。