Single-Molecule Stretching Shows Glycosylation Sets Tension in the Hyaluronan-Aggrecan Bottlebrush.

ABSTRACT: 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 ∼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.

SUBMITTER: Innes-Gold SN 

PROVIDER: S-EPMC7567978 | biostudies-literature |

REPOSITORIES: biostudies-literature