N-glycans of SREC-I (scavenger receptor expressed by endothelial cells): essential role for ligand binding, trafficking and stability.
N-glycans of SREC-I (scavenger receptor expressed by endothelial cells): essential role for ligand binding, trafficking and stability.
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DOI：
10.1093/glycob/cws010
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发表时间：
2012-05
期刊：
影响因子：
4.3
通讯作者：
M. Sano;H. Korekane;K. Ohtsubo;Y. Yamaguchi;Masaki Kato;Yukinao Shibukawa;M. Tajiri;H. Adachi;Y. Wada;M. Asahi;N. Taniguchi
中科院分区：
文献类型：
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作者：
M. Sano;H. Korekane;K. Ohtsubo;Y. Yamaguchi;Masaki Kato;Yukinao Shibukawa;M. Tajiri;H. Adachi;Y. Wada;M. Asahi;N. Taniguchi
内皮细胞表达的清道夫受体（SREC-1）介导化学修饰的脂蛋白如乙酰化低密度脂蛋白（Ac-LDL）和氧化LDL的内吞作用，并与动脉粥样硬化形成有关。我们在中国仓鼠卵巢-K1细胞中表达了重组SREC-I，并鉴定了三个潜在的糖基化位点，Asn（289），Asn（382）和Asn（393），它们都是糖基化的。为了确定N-聚糖在SREC-I中的功能，我们通过Ac-LDL的细胞内分布和细胞掺入率来表征SREC-I突变蛋白。N382 Q/N393 Q和N289 Q/N382 Q/N393 Q被隔离在内质网中，导致Ac-LDL的细胞掺入严重减少。N382 Q显示正常的细胞表面驻留和对Ac-LDL的增强的亲和力，导致Ac-LDL细胞掺入升高。这些结果表明Asn（393）的N-聚糖调节SREC-1的细胞内分选，Asn（382）的N-聚糖控制配体结合亲和力。此外，我们检测到N289 Q的胰蛋白酶敏感性增强。聚糖结构分析表明，核心岩藻糖基化双触角是所有糖基化位点的共同主要结构。此外，在Asn（289）处检测到三触角和四触角作为次要成分。还在Asn（382）和Asn（393）处检测到二等分GlcNAc。SREC-Ⅰ的结构分析和同源性建模表明，在Asn（289）处带有β1- 6 GlcNAc分支的N-聚糖保护蛋白酶的攻击，从而赋予SREC-Ⅰ更高的稳定性。这些数据表明，SREC-I的Asn（289）-、Asn（382）-和Asn（393）-连接的N-聚糖在调节蛋白水解抗性、配体结合亲和力和亚细胞定位方面具有不同的功能，所有这些功能都可能参与动脉粥样硬化的发生。
Scavenger receptor expressed by endothelial cells (SREC-I) mediates the endocytosis of chemically modified lipoproteins such as acetylated low-density lipoprotein (Ac-LDL) and oxidized LDL and is implicated in atherogenesis. We produced recombinant SREC-I in Chinese hamster ovary-K1 cells and identified three potential glycosylation sites, Asn(289), Asn(382) and Asn(393), which were all glycosylated. To determine the function of N-glycans in SREC-I, we characterized SREC-I mutant proteins by intracellular distribution and the cellular incorporation rate of Ac-LDL. N382Q/N393Q and N289Q/N382Q/N393Q were sequestered in the endoplasmic reticulum, resulting in a severe reduction in the cellular incorporation of Ac-LDL. N382Q showed a normal cell surface residency and an enhanced affinity for Ac-LDL, resulting in an elevated Ac-LDL cellular incorporation. These results indicate that the N-glycan of Asn(393) regulates the intracellular sorting of SREC-I and that the N-glycan of Asn(382) controls ligand-binding affinity. Furthermore, we detected an enhanced trypsin sensitivity of the N289Q. Glycan structure analyses revealed that the core-fucosylated bi-antennary is the common major structure at all glycosylation sites. In addition, tri- and tetra-antennary were detected as minor constituents at Asn(289). A bisecting GlcNAc was also detected at Asn(382) and Asn(393). Structural analyses and homology modeling of SREC-I suggest that the N-glycan bearing a β1-6GlcNAc branch at Asn(289) protects from proteinase attack and thus confers a higher stability on SREC-I. These data indicate that Asn(289)-, Asn(382)- and Asn(393)-linked N-glycans of SREC-I have distinct functions in regulating proteolytic resistance, ligand-binding affinity and subcellular localization, all of which might be involved in the development of atherogenesis.