Core-2O-glycans are required for galectin-3 interaction with the osteoarthritis related protein lubricin

Abstract

Author contributions: SA planned and executed the glycomic analysis, KT and LA planned and executed the glycoproteomic analysis, SH performed ELISA analysis, SR performed SPR analysis, YM and JH designed the recombinant expression and experiments executed by YM. OR, LB, RK, TS and TE were responsible for control and patient sample collection. TS and GJ contributed with design and provision of rhPRG4 for the analysis, MS and AK participated in planning and supervision of galectin-3 experiments. SF, KT and NK performed the writing with input from all contributors. NK coordinated and directed the research. All authors signed off on the final version.ABSTRACT Synovial fluid lubricin (proteoglycan 4) is a mucin-type O-linked glycosylated (60% of the mass) biological lubricant involved in osteoarthritis (OA) development. Lubricin has been reported to be cross-linked by synovial galectin-3 on the lubricating articular surface. Here, we confirm that binding to galectin-3 depended on core-2 O-linked glycans, where surface plasmon resonance of a recombinant lubricin (rhPRG4) devoid of core-2 structures lacked binding capacity to recombinant galectin-3. Both galectin-3 levels and interactions with synovial lubricin were found to be decreased in late-stage OA patients coinciding with an increase of truncated and less sialylated core 1 O-glycans. These data suggest a defect cross-linking of surface active molecules in OA and provides novel insights into OA molecular pathology.

Rapid Point-of-Care Electrochemical Sensor for the Detection of Cancer Tn Antigen Carbohydrate in Whole Unprocessed Blood

“…To understand how the LUB brush stiffens mechanically from the binding of MB-PNA, consider that the structure of PNA is composed of four Tn antigen binding sub-unit domains . Likewise, the mucin domain of recombinant LUB, which is synthesized in tumor-derived Chinese hamster ovarian cells, is rich in core 1-type O-glycans that include Tn (and T-, sTn-, and Ts-antigen) . The capacity of PNA to bind multiple Tn antigens and the multiplicity of Tn antigen sites on the LUB mucin domain lead PNA to cross-link the LUB brush.…”

“…LUB presents largely as a freely jointed chain dominated by a ∼200 nm long, highly glycosylated mucin domain that lacks any secondary or tertiary structures. The recombinant LUB used in this study possesses a mucin domain glycosylated predominantly by small, O-linked sialylated core-1 glycans. , Connecting each side of the central mucin domain are two globular end domains, which presents sub-domains comparable to two globular proteins, hemopexin, which binds hemoglobin, and somatomedin B, a structural domain of vitronectin that plays a role in cell adhesion to the extra-cellular matrix . LUB’s end domains are strongly adhesive and consequently capable of adhering onto different surfaces, including metals, conducting polymers, and carbon-based substrates .…”

A Mucin-Deficient Ocular Surface Mimetic Platform for Interrogating Drug Effects on Biolubrication, Antiadhesion Properties, and Barrier Functionality

“…In contrast to lubricin’s nonmonotonic antiadhesion properties on mucin-deficient corneal epithelial cells, dye penetrance was reduced as solution concentration increased for each lubricin variant (Figure ). The association between dye uptake and lubricin concentration is perhaps unsurprising as the presence of an oligosaccharide lattice crosslinked by galectin-3 protects against rose bengal penetrance. ,, Because galectin-3, a 35-kDa β-galactoside-binding lectin, binds lubricin O -glycans expected to be accessible on both fragments and aggregates, , we anticipated that lubricin conformation would not play a dominant role in imparting barrier functionality. Consistent with this hypothesis, the rose bengal excluded area was strongly correlated with lubricin surface coverage independent of surface conformation, with a nonparametric Spearman correlation coefficient of 0.93 ( p < 0.0001) (Figure S8A).…”

Cellular Interactions with Lubricin and Hyaluronic Acid–Lubricin Composite Coatings on Gold Electrodes in Passive and Electrically Stimulated Environments

“…Instead of repeating these characterizations, we direct the interested readers to where specific information about LUB’s properties and chemistry can be found. A comprehensive characterization of the N-LUB and R-LUB glycomes is contained in refs and respectively. Characterization of N-LUB self-assembly, surface density, and protein adhesion resistance on Au and chemically modified Au surfaces is described in ref .…”