Excretion of X-prolyl dipeptidyl-aminopeptidase in human urine as determined with a new fluorogenic substrate

X-Prolyl dipeptidyl-aminopeptidase activity was found in human urine by a sensitive fluorescence assay in which a new fluorogenic substrate, 7-glycylproline-4-methylcoumarinamide, is used. The Km value was 2.9 X 10(-4) mol/liter, and the optimum pH was 8.7 in glycine-NaOH buffer. The enzyme activity was stable at 4 degrees C for at least five days. On Sephadex G-200 column chromatography, normal human urine showed a main peak with an approximate relative molecular mass of 400 000. The procedure is simple, rapid, and accurate. The enzyme activity in urine of normal adults was: 4.30 +/- 0.13 (SE) (range, approximately 1.84-8.96) micronmol/min per gram of creatinine, and 2.16 +/- 0.09 (SE) (range, approximately 0.38-6.98) micronmol/min per liter of urine.