Koller F, Hoffmann-Ostenhof O
Hoppe Seylers Z Physiol Chem. 1979 Apr;360(4):507-13. doi: 10.1515/bchm2.1979.360.1.507.
Using the technique of affinity chromatography on a myo-inositol-substituted Sepharose, the myo-inositol oxygenase from rat kidneys was purified to homogeneity. The active enzyme contains iron, most probably in its divalent form. Electrophoresis on polyacrylamide gel containing sodium dodecylsulphate causes the cleavage of the enzyme protein into apparently identical subunits with a molecular weight of approximately 17,000. The smallest active unit consists of 4 subunits, and is in a pH-dependent equilibium with species consisting of 8, 12, and 16 subunits, respectively, which all show the same specific enzyme activity. In the presence of oxygen the enzyme is highly unstable; at the early stages of inactivation it can be reactivated by reducing agents like NaBH4. Under anaerobic conditions or under the influence of Fe2-chelating agents, the enzyme is also inactivated; this inactivation is caused by the loss of iron and concomitant cleavage into the subunits. It can be reversed by incubation with FeSO4 in the presence of air. If myo-inositol and FeSO4 are present, the reactivation involves an oligomerization to the species with 16 subunits with the uptake of 8 gram-atoms of iron per mole of this species. The enzyme reaction follows Michaelis-Menten kinetics; the Michaelis constants are 4.5 x 10(-2)M for myo-inositol and 9.5 x 10(-6)M for oxygen.
利用肌醇取代的琼脂糖凝胶亲和层析技术，将大鼠肾脏中的肌醇加氧酶纯化至同质。活性酶含有铁，很可能是二价铁。在含有十二烷基硫酸钠的聚丙烯酰胺凝胶上进行电泳，会使酶蛋白裂解成分子量约为17,000的明显相同的亚基。最小的活性单位由4个亚基组成，并且与分别由8、12和16个亚基组成的物种处于pH依赖的平衡状态，所有这些物种都表现出相同的比酶活性。在有氧存在的情况下，该酶高度不稳定；在失活的早期阶段，它可以被诸如NaBH4的还原剂重新激活。在厌氧条件下或在Fe2+螯合剂的影响下，该酶也会失活；这种失活是由于铁的损失以及随之裂解成亚基所致。在空气中与FeSO4一起孵育可以使其逆转。如果存在肌醇和FeSO4，重新激活涉及寡聚化形成具有16个亚基的物种，每摩尔该物种摄取8克原子的铁。酶反应遵循米氏动力学；肌醇的米氏常数为4.5×10(-2)M，氧气的米氏常数为9.5×10(-6)M。
