Structural Basis for the Initiation of Glycosaminoglycan Biosynthesis by Human Xylosyltransferase 1

摘要

class="head no_bottom_margin" id="sec1title">IntroductionProteoglycans (PGs) are a diverse family of glycoproteins characterized by the presence of one or more covalently attached glycosaminoglycan (GAG) chains, which often dominate the biophysical properties and biological functions of PGs. GAGs are polymers of repeating disaccharide units consisting of a hexosamine and a uronic acid sugar; the polymer is modified by sulfation at various positions (, ). Depending on the hexosamine, GAGs are classified as either heparan sulfate (HS) or as chondroitin sulfate (CS) and dermatan sulfate (DS). PGs are present on the cell surface and in the extracellular matrix of all higher animals. They are essential components of extracellular matrices and play important roles in cell adhesion and migration, morphogen and growth factor signaling, immune regulation, and the inflammatory response. PG dysfunction is linked to many conditions with major public health implications, such as arthritis, diabetes, neurodegenerative diseases, atherosclerosis, and cancer (, , ).The GAG component of PGs is synthesized in the Golgi compartment during passage of the core protein through the secretory pathway. HS, CS, and DS are attached to serine residues on the core protein through a common GlcA-β1,3-Gal-β1,3-Gal-β1,4-Xyl-β1-O-Ser tetrasaccharide linker (, ). The linker is synthesized by four glycosyltransferases (GTs) acting sequentially (A); phosphorylation of the xylose by Fam20B is required for addition of the second galactose (, , ). The first enzyme in the pathway, peptide O-xylosyltransferase (XT, EC 2.4.2.26), catalyzes the transfer of xylose from uridine diphosphate (UDP)-α-D-xylose onto serine and thus determines the site(s) of GAG attachment on the core protein. The serine-linked xylose is in β-anomeric configuration and XT is therefore classified as an “inverting” GT (). Caenorhabditis elegans and Drosophila melanogaster have one XT (known as sqv-6 and OXT, respectively), whereas vertebrates have two isozymes, XT1 and XT2 (60% amino acid identity in humans) (, , ). Disruption of tetrasaccharide linker biosynthesis in mice by genetic ablation of the glucuronyl transferase results in embryonic lethality before the 8-cell stage (href="#bib18" rid="bib18" class=" bibr popnode">Izumikawa et al., 2010). The combined function of XT1 and XT2 is expected to be similarly essential, but double-knockout mice have not been described. Genetic screens in zebrafish and mouse have revealed a function of XT1 in chondrocyte maturation during bone development (href="#bib8" rid="bib8" class=" bibr popnode">Eames et al., 2011, href="#bib29" rid="bib29" class=" bibr popnode">Mis et al., 2014). XT2-deficient mice are viable, but develop polycystic liver and kidney disease (href="#bib6" rid="bib6" class=" bibr popnode">Condac et al., 2007). In humans, XYLT1 and XYLT2 mutations cause two rare diseases with skeletal abnormalities, Desbuquois dysplasia type 2 and spondylo-ocular syndrome, respectively (href="#bib5" rid="bib5" class=" bibr popnode">Bui et al., 2014, href="#bib31" rid="bib31" class=" bibr popnode">Munns et al., 2015). The phenotypes suggest that XT1 and XT2 are not fully redundant, consistent with their somewhat different expression patterns (href="#bib8" rid="bib8" class=" bibr popnode">Eames et al., 2011, href="#bib38" rid="bib38" class=" bibr popnode">Roch et al., 2010).href="/pmc/articles/PMC5992326/figure/fig1/" target="figure" rid-figpopup="fig1" rid-ob="ob-fig1">class="inline_block ts_canvas" href="/core/lw/2.0/html/tileshop_pmc/tileshop_pmc_inline.html?title=Click%20on%20image%20to%20zoom&p=PMC3&id=5992326_gr1.jpg" target="tileshopwindow">target="object" href="/pmc/articles/PMC5992326/figure/fig1/?report=objectonly">Open in a separate windowclass="figpopup" href="/pmc/articles/PMC5992326/figure/fig1/" target="figure" rid-figpopup="fig1" rid-ob="ob-fig1">Figure 1Crystal Structure of XT1 Complexed with UDP-Xylose and a Bikunin-Derived Acceptor Peptide(A) Schematic structure of the GAG tetrasaccharide linker. GTs involved in linker biosynthesis are indicated in red. The corresponding gene names are XYLT1 (XT1), XYLT2 (XT2), B4GALT7 (GalT1), B3GALT6 (GalT2), and B3GAT3 (GlcAT1).(B) Crystal structure of human XT1, colored from N terminus (blue) to C terminus (red). UDP-xylose (silver) and peptide >2 (pink) are shown in stick representation, as are the disulfide bonds. See also href="#mmc1" rid="mmc1" class=" supplementary-material">Figure S1.