Key Laboratory of Dairy Science of Education Ministry, Northeast Agricultural University, Harbin 150030, Heilongjiang, China.
Molecules. 2012 Dec 27;18(1):263-75. doi: 10.3390/molecules18010263.
L-Lysine (L-Lys) is an essential amino acid that plays fundamental roles in protein synthesis. Many nuclear phosphorylated proteins such as Stat5 and mTOR regulate milk protein synthesis. However, the details of milk protein synthesis control at the transcript and translational levels are not well known. In this current study, a two-dimensional gel electrophoresis (2-DE)/MS-based proteomic technology was used to identify phosphoproteins responsible for milk protein synthesis in dairy cow mammary epithelial cells (DCMECs). The effect of L-Lys on DCMECs was analyzed by CASY technology and reversed phase high performance liquid chromatography (RP-HPLC). The results showed that cell proliferation ability and β-casein expression were enhanced in DCMECs treated with L-Lys. By phosphoproteomics analysis, six proteins, including MAPK1, were identified up-expressed in DCMECs treated with 1.2 mM L-Lys for 24 h, and were verified by quantitative real-time PCR (qRT-PCR) and western blot. Overexpression and siRNA inhibition of MAPK1 experiments showed that MAPK1 upregulated milk protein synthesis through Stat5 and mTOR pathway. These findings that MAPK1 involves in regulation of milk synthesis shed new insights for understanding the mechanisms of milk protein synthesis.
L-赖氨酸（L-Lys）是一种必需氨基酸，在蛋白质合成中起着重要作用。许多核磷酸化蛋白，如 Stat5 和 mTOR，调节乳蛋白合成。然而，乳蛋白合成在转录和翻译水平上的调控细节尚不清楚。在本研究中，采用二维凝胶电泳（2-DE）/MS 基于蛋白质组学技术鉴定参与奶牛乳腺上皮细胞（DCMECs）乳蛋白合成的磷酸蛋白。通过 CASY 技术和反相高效液相色谱（RP-HPLC）分析 L-Lys 对 DCMECs 的影响。结果表明，用 L-Lys 处理的 DCMECs 细胞增殖能力和β-酪蛋白表达增强。通过磷酸蛋白质组学分析，在 1.2mM L-Lys 处理 24 小时的 DCMECs 中鉴定出 6 种蛋白（包括 MAPK1）表达上调，并通过定量实时 PCR（qRT-PCR）和 Western blot 验证。MAPK1 的过表达和 siRNA 抑制实验表明，MAPK1 通过 Stat5 和 mTOR 途径上调乳蛋白合成。这些 MAPK1 参与乳合成调控的发现为理解乳蛋白合成的机制提供了新的见解。
Zotero 插件
