Ramakrishna S, Pucci D L, Benjamin W B
J Biol Chem. 1983 Apr 25;258(8):4950-6.
ATP-citrate lyase from rat liver and adipose tissue is phosphorylated by either ATP-citrate lyase kinase or catalytic subunit of cyclic AMP-dependent protein kinase to 0.5-0.6 mol/subunit. We previously demonstrated that the site phosphorylated by ATP-citrate lyase kinase (peptide B) is different from that phosphorylated by catalytic subunit of cyclic AMP-dependent protein kinase (peptide A) (Ramakrishna, S., Pucci, D. L., and Benjamin, W.B. (1981) J. Biol. Chem. 256, 10213-10216). ATP-citrate lyase phosphorylation by both protein kinases added simultaneously was increased synergistically. When ATP-citrate lyase was first phosphorylated by catalytic subunit of cyclic AMP-dependent protein kinase, the net phosphorylation of the fragments subsequently phosphorylated by lyase kinase increased about 6-fold. However, when ATP-citrate lyase was first phosphorylated by lyase kinase, there was no effect on the subsequent phosphorylation of the enzyme by cyclic AMP-dependent protein kinase. Alkaline phosphatase-dephosphorylated ATP-citrate lyase was phosphorylated by catalytic subunit of cyclic AMP-dependent protein kinase to 0.9-1.0 mol/subunit. However, dephospho-ATP-citrate lyase was not phosphorylated by lyase kinase. The addition of both protein kinases simultaneously phosphorylated ATP-citrate lyase up to 2 mol/subunit. Phosphorylation of dephospho-ATP-citrate lyase first by catalytic subunit of cyclic AMP-dependent protein kinase and ATP enabled the lyase to be phosphorylated by lyase kinase. Peptide mapping and phosphoamino acid analysis of dephospho-ATP-citrate lyase phosphorylated by catalytic subunit of cyclic AMP-dependent protein kinase and/or lyase kinase conclusively showed that phosphorylation of ATP-citrate lyase by ATP-citrate lyase kinase was completely dependent on peptide A phosphorylation by cyclic AMP-dependent protein kinase. Furthermore, increased phosphorylation when both protein kinases were added simultaneously was due to increased phosphorylation at peptide B.
大鼠肝脏和脂肪组织中的ATP-柠檬酸裂解酶可被ATP-柠檬酸裂解酶激酶或环磷酸腺苷依赖性蛋白激酶的催化亚基磷酸化，磷酸化程度为0.5 - 0.6摩尔/亚基。我们之前证明，被ATP-柠檬酸裂解酶激酶磷酸化的位点（肽段B）与被环磷酸腺苷依赖性蛋白激酶催化亚基磷酸化的位点（肽段A）不同（拉马克里希纳，S.，普奇，D. L.，和本杰明，W. B.（1981年）《生物化学杂志》256，10213 - 10216）。同时添加两种蛋白激酶对ATP-柠檬酸裂解酶的磷酸化作用具有协同增强效果。当ATP-柠檬酸裂解酶首先被环磷酸腺苷依赖性蛋白激酶的催化亚基磷酸化后，随后被裂解酶激酶磷酸化的片段的净磷酸化程度增加了约6倍。然而，当ATP-柠檬酸裂解酶首先被裂解酶激酶磷酸化时，对随后环磷酸腺苷依赖性蛋白激酶对该酶的磷酸化没有影响。碱性磷酸酶去磷酸化的ATP-柠檬酸裂解酶被环磷酸腺苷依赖性蛋白激酶的催化亚基磷酸化至0.9 - 1.0摩尔/亚基。然而，去磷酸化的ATP-柠檬酸裂解酶不能被裂解酶激酶磷酸化。同时添加两种蛋白激酶可将ATP-柠檬酸裂解酶磷酸化至2摩尔/亚基。首先由环磷酸腺苷依赖性蛋白激酶的催化亚基和ATP对去磷酸化的ATP-柠檬酸裂解酶进行磷酸化，能使该裂解酶被裂解酶激酶磷酸化。对被环磷酸腺苷依赖性蛋白激酶的催化亚基和/或裂解酶激酶磷酸化的去磷酸化ATP-柠檬酸裂解酶进行肽图谱分析和磷酸氨基酸分析，最终表明ATP-柠檬酸裂解酶激酶对ATP-柠檬酸裂解酶的磷酸化完全依赖于环磷酸腺苷依赖性蛋白激酶对肽段A的磷酸化。此外，同时添加两种蛋白激酶时磷酸化作用增强是由于肽段B的磷酸化增加所致。
