猪肌肉碳酸酐酶III的纯化及性质

Purification and properties of pig muscle carbonic anhydrase III.

作者信息

Pullan L M, Noltmann E A

出版信息

Biochim Biophys Acta. 1985 Apr 17;839(2):147-54. doi: 10.1016/0304-4165(85)90031-5.

Abstract

Pig muscle carbonic anhydrase III (carbonate hydro-lyase, EC 4.2.1.1) has been isolated and purified to homogeneity with chromatographic techniques. It has been found to be a 30 kDa protein displaying the same three activities (CO2 hydratase, acetate esterase, p-nitrophenyl phosphatase) previously described for the rabbit muscle isoenzyme, including the phosphatase activity not seen in the erythrocyte isoenzymes. The turnover numbers of the three activities are of the same order of magnitude as previously reported for rabbit muscle carbonic anhydrase III. Km and Vmax for the pig muscle CO2 hydratase activity were found to be 83 mM and 6000 s-1, respectively. The extinction coefficient at 280 nm (1 cm light path) is 22.2 for a 1% solution. Five half-cystine residues determined by performic acid oxidation are free for reaction with p-mercuribenzoate but only four are accessible to titration with dithiobisnitrobenzene. The amino acid composition of the pig muscle isoenzyme III has a high level of homology compared with that of rabbit and bovine muscle carbonic anhydrases III.

摘要

猪肌肉碳酸酐酶III（碳酸水解酶，EC 4.2.1.1）已通过色谱技术分离纯化至同质。已发现它是一种30 kDa的蛋白质，具有先前针对兔肌肉同工酶描述的相同三种活性（CO2水合酶、乙酸酯酶、对硝基苯磷酸酶），包括红细胞同工酶中未见到的磷酸酶活性。这三种活性的周转数与先前报道的兔肌肉碳酸酐酶III处于同一数量级。发现猪肌肉CO2水合酶活性的Km和Vmax分别为83 mM和6000 s-1。对于1%的溶液，280 nm（1 cm光程）处的消光系数为22.2。通过过甲酸氧化确定的五个半胱氨酸残基可自由与对汞苯甲酸反应，但只有四个可被二硫代双硝基苯滴定。与兔和牛肌肉碳酸酐酶III相比，猪肌肉同工酶III的氨基酸组成具有高度同源性。