Ozegowski J H, Müller P J
Zentralbl Bakteriol Mikrobiol Hyg A. 1984 Dec;258(2-3):159-72.
Acid phosphatase was purified from culture filtrates of Streptomyces hygroscopicus strain JA 6599-R 27/158. Method used included as first step either ammonium sulfate precipitation or adsorption of acid phosphatase on Bentonit and the desorption of enzyme from Bentonit with alkaline buffers, adsorption to DEAE-cellulose, column chromatography on Sephadex G 50 and isoelectric focusing in Sephadex gel. The specific activity of the resulting enzyme was 51 muMol/min/mg at 25 degrees C and pH of 6.25 with p-nitrophenylphosphate as substrate. The pI detected by isoelectric focusing was at pH 7.25. The molecular weight determined by gel chromatography and by SDS electrophoresis was found to be 27 000. The pH-dependence of hydrolytic activity of acid phosphatase was substrate specific. The enzyme was found to hydrolyze essentially at pH 6.2 phosphoenolpyruvate, ATP, ADP, fructose-1,6-diphosphate and tyrosine-O-phosphate. The activity was inhibited by phosphate, molybdate, arsenate, vanadate, pyrophosphate and tetraborate. In the culture medium the acid phosphatase caused the release of phosphate from solid and soluted substrates. Therefore the involvement of acid phosphatase in the regulation of secondary metabolism was discussed.
酸性磷酸酶是从吸水链霉菌JA 6599-R 27/158的培养滤液中纯化得到的。所采用的方法第一步包括硫酸铵沉淀或酸性磷酸酶在膨润土上的吸附以及用碱性缓冲液从膨润土上解吸酶、吸附到DEAE-纤维素上、在Sephadex G 50上进行柱色谱分离以及在Sephadex凝胶中进行等电聚焦。以对硝基苯磷酸为底物时，所得酶在25℃和pH 6.25条件下的比活性为51 μmol/min/mg。通过等电聚焦检测到的pI为pH 7.25。通过凝胶色谱法和SDS电泳法测定的分子量为27000。酸性磷酸酶水解活性的pH依赖性具有底物特异性。发现该酶基本上在pH 6.2时水解磷酸烯醇丙酮酸、ATP、ADP、果糖-1,6-二磷酸和酪氨酸-O-磷酸。其活性受到磷酸盐、钼酸盐、砷酸盐、钒酸盐、焦磷酸盐和四硼酸盐的抑制。在培养基中，酸性磷酸酶导致固体和溶解底物中磷酸盐的释放。因此，讨论了酸性磷酸酶在次级代谢调节中的作用。
