Single-Molecule Stretching Shows Glycosylation Sets Tension in the Hyaluronan-Aggrecan Bottlebrush
Single-Molecule Stretching Shows Glycosylation Sets Tension in the Hyaluronan-Aggrecan Bottlebrush
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DOI：
10.1016/j.bpj.2020.08.016
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发表时间：
2020-10-06
影响因子：
3.4
通讯作者：
Saleh, Omar A.
中科院分区：
文献类型：
--
作者：
Innes-Gold, Sarah N.;Berezney, John P.;Saleh, Omar A.
由多糖透明质酸（HA）和蛋白聚糖聚集蛋白聚糖形成的大瓶刷复合物有助于软骨抗压性，并且是健康关节功能所必需的。各种机械力作用于软骨细胞外基质中的这些复合物，激发了对将其结构和机械响应联系起来的定量描述的需要。使用电子显微镜的研究已经成像HA-聚集蛋白聚糖刷，但需要吸附到表面，显着改变其天然构象的复合物。我们使用磁镊力谱测量的变化，在扩展和机械响应的HA链聚集蛋白聚糖单体结合，形成一个bottlebrush。该技术直接测量单个复合物随时间推移和在不同溶液条件下所经历的变化。在添加聚集蛋白聚糖后，我们发现当HA链处于非常低的外部张力下（即，拉伸力小于1 pN）。我们使用模型的力延伸行为表明，聚集蛋白聚糖之间的排斥引起的HA链的内部张力。通过参考瓶刷聚合物行为的理论，我们证明了内部张力的实验值是一致的多分散聚集蛋白聚糖人口，可能是由不同程度的糖基化。通过酶促去糖基化的聚集蛋白聚糖，我们表明，聚集蛋白聚糖糖基化是导致HA硬化的结构特征。然后，我们构建了一个简单的随机结合模型，表明可变的糖基化导致HA内部张力分布广泛，在更长的长度尺度上引起力学变化。我们的研究结果提供了一个机制图片的灵活性和大小的HA和聚集蛋白聚糖导致刷架构和机械性能的这一重要组成部分的软骨。
Large bottlebrush complexes formed from the polysaccharide hyaluronan (HA) and the proteoglycan aggrecan contribute to cartilage compression resistance and are necessary for healthy joint function. A variety of mechanical forces act on these complexes in the cartilage extracellular matrix, motivating the need for a quantitative description that links their structure and mechanical response. Studies using electron microscopy have imaged the HA-aggrecan brush but require adsorption to a surface, dramatically altering the complex from its native conformation. We use magnetic tweezers force spectroscopy to measure changes in extension and mechanical response of an HA chain as aggrecan monomers bind and form a bottlebrush. This technique directly measures changes undergone by a single complex with time and under varying solution conditions. Upon addition of aggrecan, we find a large swelling effect manifests when the HA chain is under very low external tension (i.e., stretching forces less than similar to 1 pN). We use models of force-extension behavior to show that repulsion between the aggrecans induces an internal tension in the HA chain. Through reference to theories of bottlebrush polymer behavior, we demonstrate that the experimental values of internal tension are consistent with a polydisperse aggrecan population, likely caused by varying degrees of glycosylation. By enzymatically deglycosylating the aggrecan, we show that aggrecan glycosylation is the structural feature that causes HA stiffening. We then construct a simple stochastic binding model to show that variable glycosylation leads to a wide distribution of internal tensions in HA, causing variations in the mechanics at much longer length scales. Our results provide a mechanistic picture of how flexibility and size of HA and aggrecan lead to the brush architecture and mechanical properties of this important component of cartilage.