Integumental phosphatase isoenzymes from white puparia of Ceratitis capitata: isolation and characterization.
Two specific alkaline phosphatase forms were identified in the integument of wild-type Ceratitis capitata during transition of larvae to pupae. The separation was achieved by DEAE-cellulose chromatography; alkaline phosphatase 1 and alkaline phosphatase 2 were eluted in 0.1 and 0.4 M KCl, respectively. Both isoenzymes have a molecular weight of approximately 180,000. The pH curve reveals two peaks for both alkaline phosphatases: one at 9.4 and the other at 11.0. The two isoenzymes at both pH optima catalyze the hydrolysis of phosphotyrosine and beta-glycerophosphate, but not phosphoserine, phosphothreonine, ATP, or AMP. However, at pH 9.4, alkaline phosphatase 1 is more effective than ALPase 2 and exhibits a preference for phosphotyrosine. The divalent cations Mn2+, Mg2+, and Ba2+ activate the enzymes, while Cu2+ and Zn2+ are inhibitors for both isoenzymes. Both isoenzymes are inactivated by EDTA. The effect of amino acids on enzyme activity was also tested. Alkaline phosphatase 1 is inhibited by L-tyrosine, while alkaline phosphatase 2 is unaffected. L-Phenylalanine has no effect on either isoenzyme. Both isoenzymes are inhibited by urea and 2-mercaptoethanol. Simultaneous addition of urea and 2-mercaptoethanol reveals that ALPase 1 is more sensitive to these inhibitors than ALPase 2.
Bourtzis
K
K
Department of Biology, University of Patras, Greece.
Marmaras
V J
VJ
D000818:Animals; D004175:Diptera / Q000201:enzymology*; D005994:Glycerophosphates / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007527:Isoenzymes; D007814:Larva / Q000201:enzymology; D009024:Morphogenesis; D009596:Nitrophenols / Q000378:metabolism; D009943:Organophosphorus Compounds / Q000378:metabolism; D010744:Phosphoric Monoester Hydrolases / Q000032:analysis* / Q000302:isolation & purification; D019000:Phosphotyrosine; D011679:Pupa / Q000201:enzymology; D013347:Subcellular Fractions / Q000201:enzymology; D014443:Tyrosine / Q000031:analogs & derivatives / Q000378:metabolism
D005994:Glycerophosphates; D007527:Isoenzymes; D009596:Nitrophenols; D009943:Organophosphorus Compounds; D019000:Phosphotyrosine; C008644:nitrophenylphosphate; D014443:Tyrosine; D010744:Phosphoric Monoester Hydrolases; C031463:beta-glycerophosphoric acid