Allantoinase catalyzes the hydrolysis of allantoin to allantoic acid, a reaction important in both biogenesis and degradation of ureides. Ureide production in cotyledons of germinating soybean (Glycine max L.) seeds has not been studied extensively but may be important in mobilizing nitrogen reserves. Allantoinase was purified approximately 2500-fold from a crude extract of soybean seeds by differential centrifugation, heat treatment, ammonium sulfate fractionation, ethanol fractionation, and fast protein liquid chromatography (Pharmacia) with Mono-Q and Superose columns. The purified enzyme had a subunit size of 30 kD. Polyclonal antibodies produced against the purified protein titrated allantoinase activity in a crude extract of seed proteins. Antibodies recognized the 30-kD band in western blot analysis of crude seed extracts, indicating that they were specific for allantoinase.
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机译:尿囊素酶催化尿囊素水解为尿囊酸,该反应对于生物发生和脲离子的降解都很重要。尚未广泛研究发芽大豆(Glycine max L.)种子子叶中的尿素生产,但对调动氮储量可能很重要。通过差速离心,热处理,硫酸铵分级分离,乙醇分级分离和具有Mono-Q和Superose柱的快速蛋白质液相色谱法(Pharmacia),从大豆种子的粗提物中纯化尿囊素酶约2500倍。纯化的酶具有30kD的亚单位大小。针对纯化蛋白产生的多克隆抗体滴定了种子蛋白粗提物中尿囊素的活性。抗体在粗种子提取物的蛋白质印迹分析中识别出30 kD条带,表明它们对尿囊素酶具有特异性。
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