Zn++ dependent acid phosphatase from chicken's liver was purified to almost homogeneity by heat treatment at 60T, cation exchange chromatopaphy on CM-Cellulose at pH 5.5, gel filtration on Sephadex G-150 and affinity chromatogmphy on Sepharosic tartramic acid affinity gel with specific activity of 4U/mg and recovery of 8%. Purification achieved was 50 fold. He purified enzyme migrated a single band. The enzyme ha molecular weight of 100,000 and is a dimer with two apparent identical subunits of 48,000 - 50,000 as determined by moditan dodecyl sulphate polyacrylamide gel electrophoresis. The enzyme requires Zn++ for catalytic activity. The maximum enzyme activity we obtained in the psesence of 5 - 8 mM ZnCl2 at pH 6.0. Higher concentrations of Zn++ were found to be inhibitory. As compound to Za++ activation, Mn++ and Co++ showed 32% and 16% activation respectively. The enzyme had optimum pH 5.5 - 7.5, optimum temperature 55°C and was stable at 40°C. The enzyme showed restricted specificity. p nitrephenylphosphate we found to be a better abstract while phenylphosphate, 8-glyccrophosphate and FMN were also hydrolyzed at a significant rate. Other substrates like phosphoserine, ATP and phosphoenol pyruvate etc. were not hydrolyzed. NaF, a strong inhibitor of high molecular weight acid phosphatase, had no effect on Za++- dependent acid phosphates activity. However, the enzyme was inhibited by phosphate, titrate, EDTA and ATP. The Km value against p- nitrophenylphosphate as a substrate, was found to be ImM at pH 6.0. Phosphste and tartrate were competitive inhibitors with Ki 0.5mM and 6mM respectively. ATP was noncompetitive inhibitor (Ki = 2.3mM) while EDTA was found to be uncompetitive inhibitor (Ki =2.5mM).
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