Masuda Hiromi, Takenaka Yasuhiro, Shikamoto Yasuo, Kagawa Masayuki, Mizuno Hiroshi, Tsuji Frederick I
Department of Biochemistry, National Institute of Agrobiological Sciences, 2-1-2 Kannondai, Tsukuba, Ibaraki 305-8602, Japan.
Protein Expr Purif. 2003 Oct;31(2):181-7. doi: 10.1016/s1046-5928(03)00186-4.
Gradient elution chromatography of recombinant apoaequorin carried out in the presence of Ca2+ revealed two isoforms of apoaequorin, reduced and oxidized, whereas in the presence of EDTA 3 isoforms were observed. In a regeneration mixture of apoaequorin, coelenterazine, EDTA, and 2-mercaptoethanol, four isoforms were obtained, of which only one, aequorin, gave light with Ca2+. A method is described for the preparation of highly pure aequorin. The aequorin was stable in solution for approximately 10 days at 4 degrees C and pH 7.6, and then it gradually lost activity with a half-life of about 20 days until it was almost completely inactive on day 30.
在Ca2+存在的情况下对重组脱辅基水母发光蛋白进行梯度洗脱色谱分析，发现了还原型和氧化型两种脱辅基水母发光蛋白异构体，而在EDTA存在的情况下观察到了3种异构体。在脱辅基水母发光蛋白、腔肠素、EDTA和2-巯基乙醇的再生混合物中，得到了4种异构体，其中只有一种，即水母发光蛋白，能与Ca2+反应发光。描述了一种制备高纯水母发光蛋白的方法。水母发光蛋白在4℃和pH 7.6的溶液中可稳定约10天，然后逐渐失去活性，半衰期约为20天，直到第30天几乎完全失活。
