Sugihara A, Shimada Y, Nagao T, Iizumi T, Nakamura K, Fukase T, Tominaga Y
Osaka Municipal Technical Research Institute, Japan.
Biosci Biotechnol Biochem. 1994 Apr;58(4):752-5. doi: 10.1271/bbb.58.752.
An intracellular carboxylesterase from Pseudomonas sp. was overproduced in E. coli, and purified to homogeneity by a combination of hydrogen bond chromatography, gel filtration, and hydrophobic interaction chromatography. Gel filtration and SDS-PAGE suggested that the purified enzyme consisted of two subunits of molecular mass of 28 kDa. Its isoelectric point was 5.9. The enzyme was thermolabile, and showed its maximum activity at 22 degrees C (pH 7.5). Methyl propionate was hydrolyzed at the highest rate among the fatty acid methyl esters tested. PMSF, DFP, PCMB, and HgCl2 inhibited the enzyme markedly, suggesting that serine and/or cysteine is in or near the active site.
一种来自假单胞菌属的细胞内羧酸酯酶在大肠杆菌中过量表达，并通过氢键色谱、凝胶过滤和疏水相互作用色谱相结合的方法纯化至同质。凝胶过滤和SDS-PAGE表明纯化后的酶由两个分子量为28 kDa的亚基组成。其等电点为5.9。该酶对热不稳定，在22℃（pH 7.5）时表现出最大活性。在测试的脂肪酸甲酯中，丙酸甲酯的水解速率最高。PMSF、DFP、PCMB和HgCl2对该酶有明显抑制作用，表明丝氨酸和/或半胱氨酸位于活性位点或其附近。
