豚鼠胸腺中碱性磷酸酶的生化特性

Biochemical characterization of alkaline phosphatase in guinea pig thymus.

Fräki J, Ruuskanen O, Kouvalainen K

Biochim Biophys Acta. 1977 Jun 10;482(2):370-8. doi: 10.1016/0005-2744(77)90250-9.

Abstract

Alkaline phosphatase (orthophosphoric-monoester phosphohydrolase (alkaline optimum), EC 3.1.3.1) in guinea pig thymus was extracted optimally in 10 mM Tris - HCl buffer at pH 8.0 containing 5 g/l Triton X-100. 2. alpha-Glycerophosphate, beta-glycerophosphate and phenolphthalein monophosphate were hydrolyzed by thymus extract with a pH optimum at 9.8-10.0, whereas p-nitrophenylphosphate and alpha-naphthylphosphate were hydrolyzed with pH optima at 10.7-10.8 and beta-naphthylphosphate at pH 11.2. P-Nitrophenylphosphate and phenolphthalein monophosphate proved to be the most suitable substrates. 3. Alkaline phosphatase was effectively inhibited by EDTA, Zn2+, histidine and urea therefore resembling the inhibition characteristics of alkaline phosphatase in the placenta and kidney, but not that in the liver and intestine, which differed markedly. 4. DEAE-cellulose chromatography and polyacrylamide disc electrophoresis revealed three enzyme peaks which did not differ in their substrate specificities and modifier characteristics. 5. Polyacrylamide disc electrophoresis of thymus, serum, placenta, kidney, liver, bone and intestine revealed no alkaline phosphatase bands definitely unique to thymus.

摘要

豚鼠胸腺中的碱性磷酸酶（正磷酸单酯磷酸水解酶（最适pH碱性），EC 3.1.3.1）在含5 g/l Triton X-100的pH 8.0的10 mM Tris - HCl缓冲液中提取效果最佳。2. α-甘油磷酸、β-甘油磷酸和酚酞单磷酸可被胸腺提取物水解，最适pH为9.8 - 10.0，而对硝基苯磷酸和α-萘基磷酸水解的最适pH为10.7 - 10.8，β-萘基磷酸水解的最适pH为11.2。对硝基苯磷酸和酚酞单磷酸被证明是最合适的底物。3. 碱性磷酸酶可被EDTA、Zn²⁺、组氨酸和尿素有效抑制，因此类似于胎盘和肾脏中碱性磷酸酶的抑制特性，但与肝脏和肠道中碱性磷酸酶的抑制特性明显不同。4. DEAE - 纤维素色谱法和聚丙烯酰胺圆盘电泳显示有三个酶峰，它们在底物特异性和修饰剂特性方面没有差异。5. 对胸腺、血清、胎盘、肾脏、肝脏、骨骼和肠道进行聚丙烯酰胺圆盘电泳，未发现绝对特属于胸腺的碱性磷酸酶条带。