Kameda K
Hokkaido Igaku Zasshi. 1984 Mar;59(2):140-7.
The substrate specificity of the acyl-CoA synthetase from E. coli was studied. The enzyme was purified by means of DEAE-Sephacel, hydroxyapatite, Sepharose 6B, and blue dextran-Sepharose 6B chromatography. The molecular weight of the purified enzyme was 47,000 estimated by Sephadex G-200 column chromatography. 45,000 was determined as the molecular weight by sodium dodecyl sulfate polyacrylamide gel electrophoresis. The purified enzyme was quite unstable, however, the high concentration of potassium phosphate increased its stability. The chaotropic ion decreased the activity of purified enzyme, and the addition of the antichaotropic ions restored the activity to the original level. The purified enzyme activated the fatty acids with chain length of 6 to 18 carbon atoms, which was similar to the observations for the acyl-CoA synthetase in the crude extracts of E. coli. The purified enzyme also activated trans fatty acids at the same conversion rates as the corresponding cis isomers. Throughout the purification procedure, decanoyl-CoA synthetase activity was observed in the fractions which contained oleoyl-CoA synthetase activity.
对大肠杆菌酰基辅酶A合成酶的底物特异性进行了研究。该酶通过DEAE-葡聚糖凝胶、羟基磷灰石、琼脂糖6B和蓝色葡聚糖-琼脂糖6B色谱法进行纯化。通过葡聚糖凝胶G-200柱色谱法估计纯化酶的分子量为47,000。通过十二烷基硫酸钠聚丙烯酰胺凝胶电泳测定分子量为45,000。然而，纯化酶相当不稳定，高浓度的磷酸钾可提高其稳定性。离液序列高的离子会降低纯化酶的活性，而添加抗离液序列高的离子可使活性恢复到原始水平。纯化酶可激活碳链长度为6至18个碳原子的脂肪酸，这与大肠杆菌粗提物中酰基辅酶A合成酶的观察结果相似。纯化酶还以与相应顺式异构体相同的转化率激活反式脂肪酸。在整个纯化过程中，在含有油酰辅酶A合成酶活性的级分中观察到癸酰辅酶A合成酶活性。
