Phosphoglycerates and protein phosphorylation: identification of a protein substrate as glucose-1,6-bisphosphate synthetase.
Phosphoglycerates and protein phosphorylation: identification of a protein substrate as glucose-1,6-bisphosphate synthetase.
复制标题
磷酸甘油酯和蛋白质磷酸化：鉴定蛋白质底物为葡萄糖-1,6-二磷酸合成酶。
DOI：
10.1111/j.1471-4159.1991.tb02028.x
复制
发表时间：
1991
影响因子：
4.7
通讯作者：
Ueda,T
中科院分区：
文献类型：
--
作者：
Morino,H;Fischer-Bovenkerk,C;Kish,PE;Ueda,T
我们之前已经报道了哺乳动物大脑中存在两个内源性蛋白质磷酸化系统，它们在3-磷酸甘油(3PG)和三磷酸腺苷(ATP)存在下被增强。我们在这里介绍了其中一个系统的研究，72 kDa蛋白的磷酸化(3PG-PP72)。该体系通过硫酸铵分级沉淀、羟基磷灰石层析和疏水作用高效液相色谱分离得到底物3PG-PP72和一种蛋白激酶。底物蛋白被[1-32P]1，3-二磷酸甘油酸酯([1-32P]1，3BPG)直接磷酸化，其Kmof为1.1 nm。在[γ-32P]ATP和3PG存在下，非放射性1，3BPG抑制32P参入。磷酸肽图和磷酸氨基酸分析表明，在3PG和ATP存在下，3PG-PP72的磷酸化位点是与[1-32P]1，3BPG相同的丝氨酸残基。此外，在3PG和ATP存在的情况下，3PG-PP72中的[32P]磷酸通过随后与葡萄糖-1-磷酸或葡萄糖-6-磷酸的孵育而被去除。3PG-PP72的层析行为与葡萄糖-1，6-二磷酸(G1，6P2)合成酶相同。根据这些观察，我们得出结论，3PG-PP72是G1，6P合成酶，它直接被1，3BPG磷酸化，1，3BPG是由3PG和ATP形成的，3PG-PP72粗品中存在3PG激酶。
We have previously reported the occurrence of two endogenous protein phosphorylation systems in mammalian brain that are enhanced in the presence of 3‐phosphoglycerate (3PG) and ATP. We present here a study of one of these systems, the phosphorylation of the 72‐kDa protein (3PG‐PP72). This system was separated into the substrate, 3PG‐PP72, and a kinase by ammonium sulfate fractionation, hydroxyapatite chromatography, and hydrophobic interaction HPLC. The substrate protein was shown to be directly phosphorylated with [1‐32P]1,3‐bisphosphoglycerate ([1‐32P]1,3BPG) with an apparentKmof 1.1 nM. Nonradioactive 1,3BPG inhibited32P incorporation in the presence of [γ‐32P]ATP and 3PG. Phosphopeptide mapping and phosphoamino acid analyses indicated that the site of phosphorylation of 3PG‐PP72observed in the presence of 3PG and ATP is a serine residue identical to that observed with [1‐32P]1,3BPG. Moreover, [32P]phosphate incorporated into 3PG‐PP72in the presence of 3PG and ATP was removed by subsequent incubation with glucose‐1‐phosphate or glucose‐6‐phosphate. Finally, 3PG‐PP72showed chromatographic behaviors identical to those of glucose‐1,6‐bisphosphate (G1,6P2) synthetase. Based upon these observations, we conclude that 3PG‐PP72is G1,6P2synthetase and that it is phosphorylated directly by 1,3BPG, which is formed from 3PG and ATP by 3PG kinase present in a crude 3PG‐PP72preparation.