Fox H L, Pham P T, Kimball S R, Jefferson L S, Lynch C J
Department of Cellular and Molecular Physiology, The Pennsylvania State University College of Medicine, Hershey, Pennsylvania 17033, USA.
Am J Physiol. 1998 Nov;275(5):C1232-8. doi: 10.1152/ajpcell.1998.275.5.C1232.
Previous studies indicated that amino acids may activate the protein kinase activity of the target of rapamycin (TOR) and thereby augment and/or mimic the effects of insulin on protein synthesis, p70(S6k) phosphorylation, and multicellular clustering in adipocytes. To identify the individual amino acids responsible for these effects, the present study focused on the TOR substrate and translational repressor 4E-BP1. A complete mixture of amino acids stimulated the phosphorylation of 4E-BP1, decreasing its association with eukaryotic initiation factor eIF-4E. Studies on subsets of amino acids and individual amino acids showed that L-leucine was the amino acid responsible for most of the effects on 4E-BP1 phosphorylation; however, the presence of other amino acids was required to observe a maximal effect. The stimulatory effect of leucine was stereospecific and not mimicked by other branched chain amino acids but was mimicked by the leucine metabolite alpha-ketoisocaproate (alpha-KIC). The effect of alpha-KIC, but not leucine, was attenuated by the transaminase inhibitor (aminooxy)acetate. The latter result indicates that the effects of alpha-KIC required its conversion to leucine. Half-maximal stimulation of 4E-BP1 phosphorylation occurred at approximately 430 microM; therefore, the response was linear within the range of circulating concentrations of leucine found in various nutritional states.
先前的研究表明，氨基酸可能激活雷帕霉素靶蛋白（TOR）的蛋白激酶活性，从而增强和/或模拟胰岛素对脂肪细胞中蛋白质合成、p70（S6k）磷酸化和多细胞聚集的作用。为了确定导致这些效应的单个氨基酸，本研究聚焦于TOR底物及翻译阻遏物4E-BP1。氨基酸的完整混合物刺激了4E-BP1的磷酸化，减少了其与真核起始因子eIF-4E的结合。对氨基酸亚组和单个氨基酸的研究表明，L-亮氨酸是对4E-BP1磷酸化产生大部分效应的氨基酸；然而，需要其他氨基酸的存在才能观察到最大效应。亮氨酸的刺激作用具有立体特异性，其他支链氨基酸无法模拟，但亮氨酸代谢物α-酮异己酸（α-KIC）可以模拟。转氨酶抑制剂（氨氧基）乙酸可减弱α-KIC而非亮氨酸的作用。后一结果表明，α-KIC的作用需要将其转化为亮氨酸。4E-BP1磷酸化的半最大刺激浓度约为430微摩尔；因此，在各种营养状态下发现的亮氨酸循环浓度范围内，反应呈线性。
