A single molecule assay to probe monovalent and multivalent bonds between hyaluronan and its key leukocyte receptor CD44 under force.
A single molecule assay to probe monovalent and multivalent bonds between hyaluronan and its key leukocyte receptor CD44 under force.
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DOI：
10.1038/srep34176
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发表时间：
2016-09-29
影响因子：
4.6
通讯作者：
Richter RP
中科院分区：
文献类型：
--
作者：
Bano F;Banerji S;Howarth M;Jackson DG;Richter RP
糖胺聚糖（Glycosaminoglycans，GAG）是一类广泛存在于细胞外的线性阴离子多糖，是细胞功能的重要外在调节因子。尽管机械刺激在细胞通讯中的重要性已得到公认，但目前只有少数单分子方法可用于研究单价和多价GAG·蛋白质键如何响应定向机械力。在这里，我们设计了这样一种方法，通过结合目的设计的表面，提供固定的GAG和受体在控制的纳米级组织与单分子力光谱（SMFS）。我们应用该方法研究了GAG聚合物透明质酸（HA）与血管内皮细胞受体CD44的相互作用。与其较低的结合亲和力相比，HA和CD 44之间的单个键在力的作用下显着抵抗断裂。多个键沿着一个单一的HA链断裂顺序和独立的负载下。我们还展示了如何强大的非共价键，这是通用的控制蛋白质和GAG固定，可以有效地用作分子锚在SMFS。因此，我们建立了一个通用的方法来分析单GAG链水平上的GAG·蛋白质相互作用的纳米力学，这提供了新的分子水平上的洞察力的作用，机械力的组装和功能的GAG丰富的细胞外基质。
Glycosaminoglycans (GAGs), a category of linear, anionic polysaccharides, are ubiquitous in the extracellular space, and important extrinsic regulators of cell function. Despite the recognized significance of mechanical stimuli in cellular communication, however, only few single molecule methods are currently available to study how monovalent and multivalent GAG·protein bonds respond to directed mechanical forces. Here, we have devised such a method, by combining purpose-designed surfaces that afford immobilization of GAGs and receptors at controlled nanoscale organizations with single molecule force spectroscopy (SMFS). We apply the method to study the interaction of the GAG polymer hyaluronan (HA) with CD44, its receptor in vascular endothelium. Individual bonds between HA and CD44 are remarkably resistant to rupture under force in comparison to their low binding affinity. Multiple bonds along a single HA chain rupture sequentially and independently under load. We also demonstrate how strong non-covalent bonds, which are versatile for controlled protein and GAG immobilization, can be effectively used as molecular anchors in SMFS. We thus establish a versatile method for analyzing the nanomechanics of GAG·protein interactions at the level of single GAG chains, which provides new molecular-level insight into the role of mechanical forces in the assembly and function of GAG-rich extracellular matrices.
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DOI：
10.1084/jem.183.3.1119
发表时间：
1996-03-01
期刊：
The Journal of experimental medicine
影响因子：
--
作者：
DeGrendele HC;Estess P;Picker LJ;Siegelman MH
通讯作者：
Siegelman MH
影响因子：
16.8
作者：
Banerji, Suneale;Wright, Alan J.;Jackson, David G.
通讯作者：
Jackson, David G.
DOI：
10.1073/pnas.061324998
发表时间：
2001-03-27
影响因子：
11.1
作者：
Evans, E;Leung, A;Simon, S
通讯作者：
Simon, S
影响因子：
3.4
作者：
Evans, E;Ritchie, K
通讯作者：
Ritchie, K
影响因子：
3.4
作者：
Christophis, Christof;Taubert, Isabel;Rosenhahn, Axel
通讯作者：
Rosenhahn, Axel