Molecular basis of collagen galactosylation by GLT25D1.

Abstract

GLT25D1 O-galactosylates hydroxylysine residues in collagen and is essential for collagen maturation and function. Dysfunctions of GLT25D1 cause various tissue disorders. Despite its biological significance, the action mechanism of GLT25D1 remains enigmatic. Here we report the cryo-EM structures of human GLT25D1 and its ternary complex with UDP and hydroxylated acceptor substrates, revealing a bi-lobe architecture for the GLT25D1 monomer that organizes into dimeric and hexameric oligomers. The N-lobe of GLT25D1 contains a high-affinity UDP-galactose binding site, and the C-lobe is the catalytic domain of the enzyme. The structures together with biochemical analyses unravel the key recognition of the consensus "Hyl-Gly" motif from collagen acceptor substrates and associated catalytic mechanism. We further demonstrate that GLT25D1 mutations linked to cerebral small vessel disease and musculoskeletal defects adversely affect its function via distinct mechanisms. Our findings elucidate the molecular mechanism underlying collagen glycosylation and provide a molecular framework for understanding GLT25D1-related diseases.