Witters L A, Watts T D, Daniels D L, Evans J L
Department of Medicine, Dartmouth Medical School, Hanover, NH 03756.
Proc Natl Acad Sci U S A. 1988 Aug;85(15):5473-7. doi: 10.1073/pnas.85.15.5473.
The mechanism underlying the ability of insulin to acutely activate acetyl-CoA carboxylase [acetyl-CoA: carbon-dioxide ligase (ADP-forming), EC 6.4.1.2; AcCoA-Case] has been examined in Fao Reuber hepatoma cells. Insulin promotes the rapid activation of AcCoACase, as measured in cell lysates, and this stimulation persists to the same degree after isolation of AcCoACase by avidin-Sepharose chromatography. The insulin-stimulated enzyme, as compared with control enzyme, exhibits an increase in both citrate-independent and -dependent activity and a decrease in the Ka for citrate. Direct examination of the phosphorylation state of isolated 32P-labeled AcCoACase after insulin exposure reveals a marked decrease in total enzyme phosphorylation coincident with activation. The dephosphorylation due to insulin appears to be restricted to the phosphorylation sites previously shown to regulate AcCoACase activity. All of these effects of insulin are mimicked by a low molecular weight autocrine factor, tentatively identified as an oligosaccharide, present in conditioned medium of hepatoma cells. These data suggest that insulin may activate AcCoACase by inhibiting the activity of protein kinase(s) or stimulating the activity of protein phosphatase(s) that control the phosphorylation state of the enzyme.
在Fao Reuber肝癌细胞中研究了胰岛素急性激活乙酰辅酶A羧化酶[乙酰辅酶A：二氧化碳连接酶（ADP形成），EC 6.4.1.2；AcCoA-Case]能力的潜在机制。胰岛素可促进AcCoACase的快速激活，这在细胞裂解物中得到了测定，并且在用抗生物素蛋白-琼脂糖凝胶色谱法分离AcCoACase后，这种刺激仍保持相同程度。与对照酶相比，胰岛素刺激的酶在不依赖柠檬酸和依赖柠檬酸的活性方面均有所增加，并且对柠檬酸的解离常数降低。直接检测胰岛素作用后分离出的32P标记的AcCoACase的磷酸化状态，发现总酶磷酸化与激活同时显著降低。胰岛素引起的去磷酸化似乎仅限于先前显示可调节AcCoACase活性的磷酸化位点。胰岛素的所有这些作用都被一种低分子量自分泌因子模拟，该因子初步鉴定为一种寡糖，存在于肝癌细胞的条件培养基中。这些数据表明，胰岛素可能通过抑制控制该酶磷酸化状态的蛋白激酶活性或刺激蛋白磷酸酶活性来激活AcCoACase。
