Differential glycomics of epithelial membrane glycoproteins from urinary exovesicles reveals shifts towards complex-type N-glycosylation in classical galactosemia
Abstract
Point mutations in the gene encoding galactose-1-phosphate uridyltransferase (GALT) cause activity loss of the enzyme and toxic effects mediated by accumulating metabolic intermediates of galactose in untreated, non-dietary-controlled patients with classical galactosemia. However, also in patients on a lactose-free diet endogenously formed galactose mediates severe chronically toxic effects leading to clinical symptoms, like cognitive impairment and ovarian dysfunction in females. Under galactose stress the cosubstrate of GALT, galactose-1-phosphate, accumulates, and disturbs catabolic and anabolic pathways of the central carbohydrate metabolism. Enzymatic imbalances may have impact on the formation of cosubstrate involved in glycoprotein galactosylation and hence cause shifts in glycosylation patterns of membrane proteins. A suitable non-invasive material for differential glycomics derived from epithelial plasma membranes was found in urinary exovesicles and in the shed, N-glycosylated Tamm-Horsfall protein. By applying matrix-assisted laser ionization mass spectrometry we demonstrate that GALT deficiency is associated with dramatic shifts from prevalent high-mannose-type glycans found in healthy subjects towards complex-type N-linked glycosylation in patients. These N-glycosylation shifts were observed on exosomal N-glycoproteins, but not on the Tamm-Horsfall glycoprotein, which showed predominant high-mannose-type glycosylation with M6.
References
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