Identification of a pocket in the PDK1 kinase domain that interacts with PIF and the C-terminal residues of PKA
Identification of a pocket in the PDK1 kinase domain that interacts with PIF and the C-terminal residues of PKA
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DOI：
10.1093/emboj/19.5.979
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发表时间：
2000-03-01
期刊：
影响因子：
11.4
通讯作者：
Alessi, DR
中科院分区：
文献类型：
--
作者：
Biondi, RM;Cheung, PCF;Alessi, DR
3-磷酸肌醇依赖性蛋白激酶-1（PDK 1）磷酸化并激活AGC亚家族的许多蛋白激酶。PDK 1的激酶结构域通过疏水基序与蛋白激酶C相关激酶2（PRK 2）的一个区域相互作用，称为PDK 1相互作用片段（PIF）。在这里，我们鉴定了PDK 1激酶结构域的小叶中的疏水口袋，与ATP和底物结合位点分开，其与PIF相互作用，预测形成该疏水口袋的一部分的残基的突变消除或显著降低PDK 1对PIF的亲和力，PIF增加PDK 1磷酸化合成十二肽的速率（T308 tide），对应于PKB的PDK 1磷酸化位点周围的序列。该肽是PDK 1的不良底物，但包含与PIF的PDK 1结合基序融合的T308 tide的肽是PDK 1的非常优越的上级底物，我们的研究结果表明，PIF结合口袋上的激酶域的PDK 1作为一个“对接点”，使其能够相互作用，并提高其底物的磷酸化。
The 3-phosphoinositide-dependent protein kinase-1 (PDK1) phosphorylates and activates a number of protein kinases of the AGC subfamily. The kinase domain of PDK1 interacts with a region of protein kinase C-related kinase-2 (PRK2), termed the PDK1-interacting fragment (PIF), through a hydrophobic motif, Here we identify a hydrophobic pocket in the small lobe of the PDK1 kinase domain, separate from the ATP- and substrate-binding sites, that interacts with PIF, Mutation of residues predicted to form part of this hydrophobic pocket either abolished or significantly diminished the affinity of PDK1 for PIF, PIF increased the rate at which PDK1 phosphorylated a synthetic dodecapeptide (T308tide), corresponding to the sequences surrounding the PDK1 phosphorylation site of PKB, This peptide is a poor substrate for PDK1, but a peptide comprising T308tide fused to the PDK1-binding motif of PIF was a vastly superior substrate for PDK1, Our results suggest that the PIF-binding pocket on the kinase domain of PDK1 acts as a 'docking site', enabling it to interact with and enhance the phosphorylation of its substrates.