Popov V N, Igamberdiev A U, Volvenkin S V
Biokhimiia. 1996 Oct;61(10):1898-903.
Key enzymes of glyoxylate cycle, isocitrate lyase and malate synthase, are active in the fasting rat liver. The enzymes were synthesized on day 3 after food deprivation and their activities were maximal on day 5 of fasting. Specific activity of isocitrate lyase was 0.06 units/mg protein and specific activity of malate synthase was 0.03 units/mg protein. Isocitrate lyase was isolated and purified by ammonium sulfate fractionation, DEAE-cellulose chromatography and Toyopearl HW-65 gel filtration. Enzyme was purified to specific activity of 9.0 units/mg protein with 8.2% yield. Molecular mass of isocitrate lyase was 145 kD according to gel filtration. Catalytic characteristics of isocitrate lyase indicate that the enzyme follows Michaelis-Menten kinetics (Km for isocitrate is 0.07 mM), is competitively inhibited by glucose-I-phosphate (Ki = 1.1 mM) and glucose-6-phosphate (Ki = 1.9 mM), and is activate by ADP; optimal pH is 7.4. Malate synthase was partially purified by ammonium sulfate fractionation and Sephadex G-25 gel filtration. Enzyme was purified to specific activity of 0.15 units/mg protein with 45% yield. Km of malate synthase for acetyl-CoA was 0.2 mM and Km for glyoxylate was 0.3 mM; optimal pH was 7.6.
乙醛酸循环的关键酶，异柠檬酸裂解酶和苹果酸合酶，在饥饿大鼠肝脏中具有活性。这些酶在禁食后第3天合成，其活性在禁食第5天达到最大值。异柠檬酸裂解酶的比活性为0.06单位/毫克蛋白质，苹果酸合酶的比活性为0.03单位/毫克蛋白质。异柠檬酸裂解酶通过硫酸铵分级分离、DEAE-纤维素色谱和Toyopearl HW-65凝胶过滤进行分离和纯化。酶被纯化至比活性为9.0单位/毫克蛋白质，产率为8.2%。根据凝胶过滤，异柠檬酸裂解酶的分子量为145 kD。异柠檬酸裂解酶的催化特性表明该酶遵循米氏动力学（异柠檬酸的Km为0.07 mM），受到磷酸葡萄糖-1（Ki = 1.1 mM）和磷酸葡萄糖-6（Ki = 1.9 mM）的竞争性抑制，并被ADP激活；最适pH为7.4。苹果酸合酶通过硫酸铵分级分离和Sephadex G-25凝胶过滤进行部分纯化。酶被纯化至比活性为0.15单位/毫克蛋白质，产率为45%。苹果酸合酶对乙酰辅酶A的Km为0.2 mM，对乙醛酸的Km为0.3 mM；最适pH为7.6。
