摘要
用硫酸铵盐析，磷酸盐除果胶和两次ＤＥＡＥ纤维素柱层析等方法，从毛花猕猴桃（ＡｃｔｉｎｉｄｉａｅｒｉａｎｔｈａＢｅｎｔｈ．）无细胞提取液中提纯蛋白酶，在聚丙烯酰胺凝胶电泳上呈现一条带，纯酶的活力为３２５Ｕ／ｍｇ，比活力提高８９倍，总收率约为３７％。用ＳＤＳ聚丙烯酰胺凝胶电泳测定分子量为２３ｋＤ，用等电聚焦电泳测定等电点为４８。酶的最适ｐＨ为３８，最适温度为４３℃左右。纯酶制剂对其底物牛血红蛋白的Ｋｍ值为５５６×１０－３ｍｍｏｌ。酶的紫外吸收光谱最大值为２７８ｎｍ。二巯基苏糖醇、巯基乙醇、Ｌ半胱氨酸盐酸盐等还原剂对该酶有明显的激活作用，而碘乙酸、对氯汞苯甲酸对其有抑制作用，这表明该酶属于巯基酶类。
Protease of Actinidia eriantha Benth. was purified to electrophoretic homogeneity by a
combination of ammonium sulfate precipitation, removement of pectin by phosphate, two times
chromatography on DEAE cellulose. The specific activity of purified enzyme was 325 U/mg
protein, which represented as 89 fold purification with a 37% yield. The molecular weight was
estimated to be about 23 kD by the SDS polyacrylamide gel electrophoresis, and the isoelectric
point was found to be 4.8 by isoelectric focusing. The optimum pH for the enzyme was 3.8,
optimum temperature was 43℃, and its Km value for bovine hemoglobin was 5.56×10 -3
mmol. The maximum absorption of UV spectra of the enzyme was at 278 nm. Some reductants
such as DTT, ME, L cysteine stimulated the enzyme activity, while I AA, PCMB inhibited activity.
It suggested that the enzyme is a thiol protease.
出处
《植物资源与环境》
CSCD
1999年第3期1-6,共6页
Journal of Plant Resources and Environment
基金
福建省自然科学基金
