The Presence of Outer Arm Fucose Residues on the N-Glycans of Tissue Inhibitor of Metalloproteinases-1 Reduces Its Activity
The Presence of Outer Arm Fucose Residues on the N-Glycans of Tissue Inhibitor of Metalloproteinases-1 Reduces Its Activity
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DOI：
10.1021/pr400276r
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发表时间：
2013-08-01
影响因子：
4.4
通讯作者：
Lee, Seung-Taek
中科院分区：
文献类型：
--
作者：
Kim, Han Ie;Saldova, Radka;Lee, Seung-Taek
金属蛋白酶组织抑制剂-1（TIMP-1）通过以1：1的化学计量结合来抑制基质金属蛋白酶（MNIP）。在这里，我们已经显示了N-糖基化参与TIMP-1的MMP抑制能力。从过度表达TIMP-1的HEK 293细胞纯化的TIMP-1（293 TIMP-1）显示出比从用TIMP-1杆状病毒感染的成纤维细胞或SF 9昆虫细胞纯化的TIMP-1更低的对MNIP的结合和抑制能力。TIMP-1去糖基化后，所有形式的TIMP-1显示出相似的MMP结合和抑制水平，表明糖基化参与这些TMP-1活性的调节。N-聚糖结构的分析表明，SF 9 TIMP-1具有最简单的N-聚糖结构，其次是成纤维细胞TIMP-1和293 TIMP-1，其N-聚糖结构的复杂性依次增加。进一步分析显示，从293 TIMP-1的N-聚糖切割外臂岩藻糖残基或敲低FUT 4和FUT 7（其编码将外臂岩藻糖残基添加到N-聚糖的岩藻糖基转移酶）增强了293 TIMP-1的MMP结合和催化能力，使其达到其他TIMP-1的水平。这些结果表明，TIMP-1抑制MNIP的能力至少部分受其N-聚糖的外臂岩藻糖基化调节。
Tissue inhibitor of metalloproteinases-1 (TIMP-1) inhibits matrix metalloproteinases (MNIPs) by binding at a 1:1 stoichiometry. Here we have shown the involvement of N-glycosylation in the MMP inhibitory ability of TIMP-1. TIMP-1, purified from HEK 293 cells overexpressing TIMP-1 (293 TIMP-1); showed less binding and inhibitory abilities to MNIPs than TIMP-1 purified from fibroblasts or SF9 insect cells infected with TIMP-1 baculovirus. Following deglycosylation of TIMP-1, all forms of TIMP-1 showed similar levels of MMP binding and inhibition, suggesting that glycosylation is involved in the regulation of these TMP-1 activities. Analysis of the N-glycan structures showed that SF9 TIMP-1 has the simplest N-glycan structures, followed by fibroblast TIMP-1 and 293 TIMP-1, in order of increasing complexity in their N-glycan structures. Further analyses showed that cleavage of outer arm fucose residues from the N-glycans of 293 TIMP-1 or knockdown of both FUT4 and FUT7 (which encode for fucosyltransferases that add outer arm fucose residues to N-glycans) enhanced the MMP-binding and catalytic abilities of 293 TIMP-1, bringing them up to the levels of the other TIMP-1. These results demonstrate that the ability of TIMP-1 to inhibit MNIPs is at least in part regulated by outer arm fucosylation of its N-glycans.