Enzymatic cell-surface decoration with proteins using amphiphilic lipid-fused peptide substrates
Enzymatic cell-surface decoration with proteins using amphiphilic lipid-fused peptide substrates
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使用两亲性脂质融合肽底物用蛋白质进行酶促细胞表面修饰
DOI：
10.1002/chem.201900370
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发表时间：
2019
期刊：
影响因子：
--
通讯作者：
Kamiya Noriho
中科院分区：
文献类型：
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作者：
Takahara Mari;Wakabayashi Rie;Fujimoto Naoki;Minamihata Kosuke;Goto Masahiro;Kamiya Noriho
蛋白质的脂质修饰在细胞增殖等信号的激活中起着重要的作用。因此，对脂化蛋白质的需求正在上升。然而，获得高产率和高纯度的脂化蛋白质一直是具有挑战性的。我们开发了一种利用微生物谷氨酰胺转氨酶(MTG)对多种合成脂质修饰蛋白质的策略，MTG催化蛋白质中特定的谷氨酰胺(Q)和脂质融合多肽中的赖氨酸(K)之间的交联反应。合成的脂质-G3S-MRHKGS脂质(脂质：脂肪酸、生育酚、胆石酸、胆固醇)通过MTG反应成功地连接到与Q标记蛋白融合的蛋白上，得到了Q标记蛋白脂化形式的>；90 %。将纯化的脂蛋白偶联物用于体外标记细胞膜，得到了最好的胆固醇修饰锚定能力。此外，以一种简单的方式建立了细胞表面的原位修饰：将细胞置于胆固醇融合多肽、Q标记蛋白和MTG的混合物中。
Lipid modification of proteins plays a significant role in the activation of cellular signals such as proliferation. Thus, the demand for lipidated proteins is rising. However, getting a high yield and purity of lipidated proteins has been challenging. We developed a strategy for modifying proteins with a wide variety of synthetic lipids using microbial transglutaminase (MTG), which catalyzes the cross‐linking reaction between a specific glutamine (Q) in a protein and lysine (K) in the lipid‐fused peptide. The synthesized lipid‐G3S‐MRHKGS lipid (lipid: fatty acids, tocopherol, lithocholic acid, cholesterol) was successfully conjugated to a protein fused with LLQG (Q‐tagged protein) by an MTG reaction, yielding >90 % conversion of the Q‐tagged protein in a lipidated form. The purified lipid–protein conjugates were used for labeling the cell membrane in vitro, resulting in best‐anchoring ability of cholesterol modification. Furthermore, in situ cell‐surface decoration with the protein was established in a simple manner: subjection of cells to a mixture of cholesterol‐fused peptides, Q‐tagged proteins and MTG.