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Abstract
Chondroitin sulfate (CS) is a glycosaminoglycan (GAG) widely expressed on the cell surface and within the extracellular matrix, predominantly in connective tissues. Composed of alternating β1,4-N-acetylgalactosamine (GalNAc) and β1,3-glucuronic acid (GlcA) disaccharide units, CS is covalently linked to proteoglycan (PG) core proteins, such as bikunin and syndecan, via a conserved tetrasaccharide linkage region. Functionally, CS contributes to membrane flexibility, mechanical strength, tissue homeostasis, and cell signaling. Clinically, CS supplementation has demonstrated therapeutic efficacy in osteoarthritis, inflammation, and cataracts, and CS dysregulation has been implicated in neurodegenerative diseases including Parkinson’s, Alzheimer’s, and multiple sclerosis. Initially, CS biosynthesis begins with the formation of the tetrasaccharide linkage region, followed by a critical commitment step by CSGALNACT1 and CSGALNACT2 that add a preliminary GalNAc moiety. This modification distinguishes the CS pathway from heparan sulfate (HS) biosynthesis, which begins with the addition of an N-acetylglucosamine (GlcNAc) to the tetrasaccharide linkage region by EXTL3. Subsequent polymer elongation is mediated by a set of chondroitin synthase enzymes (CHPF1, CHPF2, CHSY1, and CHSY3) whose specific activities, consensus nomenclature, and overarching interactions remain incompletely understood. These enzymes belong to a diverse family of glycosyltransferases (GTs), which catalyze glycosidic bond formation across all domains of life. GTs are classified into structural fold types- GT-A, GT-B, and the less common GT-C- based on their conserved motifs, Rossmann folds, and overall architecture. Notably, CS synthases incorporate two GT-A fold domains, reflecting both their evolutionary divergence and substrate specificity. Understanding the domain architecture and functional specialization of GTs involved in CS elongation is crucial for unraveling the mechanistic nuances of GAG biosynthesis and its broader implications in health and disease.