Tailored Presentation of Carbohydrates on a Coiled Coil-Based Scaffold for Asialoglycoprotein Receptor Targeting.
Tailored Presentation of Carbohydrates on a Coiled Coil-Based Scaffold for Asialoglycoprotein Receptor Targeting.
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DOI：
10.1021/acschembio.5b00435
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发表时间：
2015-06
影响因子：
4
通讯作者：
E. Zacco;J. Hütter;Jason L. Heier;J. Mortier;P. Seeberger;B. Lepenies;B. Koksch
中科院分区：
文献类型：
--
作者：
E. Zacco;J. Hütter;Jason L. Heier;J. Mortier;P. Seeberger;B. Lepenies;B. Koksch
卷曲螺旋折叠基序代表了配体定义呈现的理想支架，因为可以将它们沿轴定位在特定距离处。我们创建了一个卷曲螺旋糖肽库来表征肝细胞上脱唾液酸糖蛋白受体（ASGPR）的碳水化合物结合位点之间的距离。糖肽文库的组成因显示的配体（半乳糖）的数量、它们在肽序列上的位置以及肽主链和碳水化合物之间的空间而异。我们确定了糖肽与肝细胞的结合，并使用流式细胞术确定了半乳糖部分与 ASGPR 相互作用的最佳距离和方向。我们通过细胞松弛素 D 的抑制研究证实了这种结合是通过 ASGPR 介导的内吞作用发生的；荧光显微镜研究显示细胞内载体肽的吸收。因此，这项研究表明卷曲螺旋基序可以用作配体合理呈递的可靠支架。
The coiled-coil folding motif represents an ideal scaffold for the defined presentation of ligands due to the possibility of positioning them at specific distances along the axis. We created a coiled-coil glycopeptide library to characterize the distances between the carbohydrate-binding sites of the asialoglycoprotein receptors (ASGPR) on hepatocytes. The components of the glycopeptide library vary for the number of displayed ligands (galactose), their position on the peptide sequence, and the space between peptide backbone and carbohydrate. We determined the binding of the glycopeptides to the hepatocytes, and we established the optimal distance and orientation of the galactose moieties for interaction with the ASGPR using flow cytometry. We confirmed that the binding occurs through endocytosis mediated by ASGPR via inhibition studies with cytochalasin D; fluorescence microscopy studies display the uptake of the carrier peptides inside the cell. Thus, this study demonstrates that the coiled-coil motif can be used as reliable scaffold for the rational presentation of ligands.