Biochemical studies on a novel vanadate- and molybdate-sensitive acid phosphatase from human epidermis.
A novel vanadate- and molybdate-sensitive human skin epidermal acid phosphatase was purified and characterized. The enzyme was extracted from epidermal sheets with a 0.1% Triton X-100 solution buffered at pH 7.0. The purification procedure consisted of molecular permeation chromatography on Sephadex G-200 followed by chromatography on hydroxylapatite using an ammonium sulfate gradient. The molecular weight of the enzyme was 82,000 and the isoelectric point was at pH 5.6. At the optimum pH (5.1) the enzyme hydrolyzed most rapidly 1-naphthyl phosphate (Km = 0.28 mM) and 4-nitrophenyl phosphate (Km = 0.28 mM). In general, the best substrates had an aromatic leaving group. Fluoride (Ki = 39 microM; noncompetitive) and phosphate (competitive) inhibited by binding to different binding sites of the enzyme. The most potent inhibitors were vanadate (Ki = 1.9 X 10(-6)M), tungstate (Ki = 1.4 X 10(-7)M), and molybdate (Ki = 2.0 X 10(-9)M). Chemical modification and kinetic experiments suggested that the activity of the enzyme is based on imidazole, tyrosyl, and carboxyl groups. Benzoyl peroxide was a relatively potent inhibitor (Ki = 5.0 X 10(-5)M; noncompetitive). This enzyme resembled the prostatic acid phosphatase with regard to substrate specificity, inhibition characteristics, and functional groups.
D000135:Acid Phosphatase / Q000302:isolation & purification / Q000378:metabolism*; D001585:Benzoyl Peroxide / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D004817:Epidermis / Q000201:enzymology*; D005260:Female; D005459:Fluorides / Q000494:pharmacology; D006651:Histocytochemistry; D006801:Humans; D008297:Male; D008982:Molybdenum / Q000494:pharmacology*; D013379:Substrate Specificity; D013644:Tartrates / Q000494:pharmacology; D014414:Tungsten / Q000494:pharmacology; D017973:Tungsten Compounds; D014638:Vanadates; D014639:Vanadium / Q000494:pharmacology*
D004791:Enzyme Inhibitors; D013644:Tartrates; D017973:Tungsten Compounds; D014639:Vanadium; C044659:molybdate; D014638:Vanadates; D008982:Molybdenum; D000135:Acid Phosphatase; D005459:Fluorides; C045951:tungstate; D014414:Tungsten; D001585:Benzoyl Peroxide