Barnett G R, Kazarinoff M N
J Biol Chem. 1984 Jan 10;259(1):179-83.
Ornithine decarboxylase (EC 4.1.1.17) has been purified 3,500-fold from the plasmodia of Physarum polycephalum. The purified material exhibited a Km for ornithine of 0.6 mM and Vmax of 20 mumol of CO2 formed per min/mg at 30 degrees C (62 mumol/min/mg at 37 degrees C). It migrated as a single protein and activity species on high pressure liquid chromatography (TSK-3000) in 0.15 M NaCl (Mr = 80,000) and in native gels containing 5, 6.5, 8, and 9.5% acrylamide. A single protein band (Mr = 43,000) was observed on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Activity was lost upon incubation with alpha-difluoromethyl[5-14C] ornithine, and the inactivated material appeared as a single Mr = 43,000 14C-band on autoradiograms of sodium dodecyl sulfate-polyacrylamide gels. The decarboxylase activity was specific for ornithine and was pyridoxal-P-dependent. The Km for pyridoxal-P (10 microM) was identical with the Kd for pyridoxal-P binding determined from the quenching of protein fluorescence (lambda ex = 282 nm, lambda em = 350 nm, maximal quenching 81%). Using specific antibody obtained from rabbit hyperimmune serum as a probe, an Mr = 43,000 immunoreactive species was detected on nitrocellulose blots of sodium dodecyl sulfate-polyacrylamide gels of plasmodial homogenates and all pooled purification fractions, but no higher molecular weight cross-reactive material was detected.
鸟氨酸脱羧酶（EC 4.1.1.17）已从多头绒泡菌的原质团中纯化了3500倍。纯化后的物质在30℃时对鸟氨酸的Km为0.6 mM，Vmax为每分钟每毫克形成20 μmol CO2（37℃时为62 μmol/min/mg）。在0.15 M NaCl（Mr = 80,000）以及含有5%、6.5%、8%和9.5%丙烯酰胺的天然凝胶中，通过高压液相色谱法（TSK - 3000）分析，它以单一的蛋白质和活性形式迁移。在十二烷基硫酸钠 - 聚丙烯酰胺凝胶电泳上观察到一条单一的蛋白质条带（Mr = 43,000）。与α - 二氟甲基[5 - 14C]鸟氨酸一起孵育后活性丧失，失活物质在十二烷基硫酸钠 - 聚丙烯酰胺凝胶的放射自显影片上呈现为一条单一的Mr = 43,000的14C条带。脱羧酶活性对鸟氨酸具有特异性，且依赖于磷酸吡哆醛。磷酸吡哆醛的Km（10 μM）与通过蛋白质荧光猝灭（激发波长λex = 282 nm，发射波长λem = 350 nm，最大猝灭81%）测定的磷酸吡哆醛结合的Kd相同。使用从兔超免疫血清获得的特异性抗体作为探针，在原质团匀浆和所有合并的纯化级分的十二烷基硫酸钠 - 聚丙烯酰胺凝胶的硝酸纤维素印迹上检测到一个Mr = 43,000的免疫反应性条带，但未检测到更高分子量的交叉反应物质。
