Chen C H, Eisenberg F
J Biol Chem. 1975 Apr 25;250(8):2963-7.
Partially purified testicular myoinositol 1-phosphate synthase was incubated with glucose 6-phosphate and NAD+. After 2 min the reaction was stopped by the addition of NaB3H4. Phosphorylated reduced sugars were isolated by ion exchange and dephosphorylated enzymatically. Scylloinositol and myoinositol, added as carriers, were re-isolated and purified to constant specific radioactivity. Since scylloinositol phosphate is uniquely related to myoinosose-2 1-phosphate, the finding of labeled scylloinositol and myoinositol is considered strong evidence for the presence of myoinosose-2 1-phosphate, an intermediate which has been postulated in the synthase-catalyzed isomerization of glucose 6-phosphate to myoinositol 1-phosphate. About one-half the amount of intermediate was demonstrable with boiled synthase, indicating firm binding of myoinosose-2 phosphate to the enzyme.
将部分纯化的睾丸肌醇1 - 磷酸合酶与6 - 磷酸葡萄糖和NAD⁺一起温育。2分钟后，通过添加NaB₃H₄终止反应。通过离子交换分离磷酸化的还原糖，并进行酶促脱磷酸化。作为载体添加的鲨肌醇和肌醇被重新分离并纯化至恒定的比放射性。由于鲨肌醇磷酸与2 - 磷酸肌醇糖独特相关，标记的鲨肌醇和肌醇的发现被认为是2 - 磷酸肌醇糖存在的有力证据，2 - 磷酸肌醇糖是在合酶催化的6 - 磷酸葡萄糖异构化为肌醇1 - 磷酸过程中假定的一种中间体。用煮沸的合酶可检测到约一半量的中间体，这表明2 - 磷酸肌醇糖与该酶紧密结合。
