Metabolism of Vertebrate Amino Sugars with N-Glycolyl Groups INCORPORATION OF N-GLYCOLYLHEXOSAMINES INTO MAMMALIAN GLYCANS BY FEEDING N-GLYCOLYLGALACTOSAMINE

被引:24
作者
Bergfeld, Anne K. [2 ]
Pearce, Oliver M. T. [2 ]
Diaz, Sandra L. [2 ]
Lawrence, Roger [2 ]
Vocadlo, David J. [3 ]
Choudhury, Biswa [2 ]
Esko, Jeffrey D. [2 ]
Varki, Ajit [1 ,2 ]
机构
[1] Univ Calif San Diego, Glycobiol Res & Training Ctr, Dept Med, La Jolla, CA 92093 USA
[2] Univ Calif San Diego, Glycobiol Res & Training Ctr, Dept Cellular & Mol Med, La Jolla, CA 92093 USA
[3] Simon Fraser Univ, Dept Mol Biol & Biochem, Burnaby, BC V5A 1S6, Canada
基金
美国国家卫生研究院;
关键词
ACETYLNEURAMINIC ACID HYDROXYLASE; NONHUMAN SIALIC-ACID; HANGANUTZIU-DEICHER ANTIGEN; GLYCOLYLNEURAMINIC ACID; MOUSE-LIVER; LINKED GLYCOSYLATION; CELL LINE; RAT-LIVER; BIOSYNTHESIS; GALNAC;
D O I
10.1074/jbc.M112.363499
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The outermost positions of mammalian cell-surface glycans are predominantly occupied by the sialic acids N-acetylneuraminic acid (Neu5Ac) and N-glycolylneuraminic acid (Neu5Gc). To date, hydroxylation of CMP-Neu5Ac resulting in the conversion into CMP-Neu5Gc is the only known enzymatic reaction in mammals to synthesize a monosaccharide carrying an N-glycolyl group. In our accompanying paper (Bergfeld, A. K., Pearce, O. M., Diaz, S. L., Pham, T., and Varki, A. (2012) J. Biol. Chem. 287, jbc.M112.363549), we report a metabolic pathway for degradation of Neu5Gc, demonstrating that N-acetylhexosamine pathways are tolerant toward the N-glycolyl substituent of Neu5Gc breakdown products. In this study, we show that exogenously added N-glycolylgalactosamine (GalNGc) serves as a precursor for Neu5Gc de novo biosynthesis, potentially involving seven distinct mammalian enzymes. Following the GalNAc salvage pathway, UDP-GalNGc is epimerized to UDP-GlcNGc, which might compete with the endogenous UDP-GlcNAc for the sialic acid biosynthetic pathway. Using UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase-deficient cells, we confirm that conversion of GalNGc into Neu5Gc depends on this key enzyme of sialic acid biosynthesis. Furthermore, we demonstrate by mass spectrometry that the metabolic intermediates UDP-GalNGc and UDP-GlcNGc serve as substrates for assembly of most major classes of cellular glycans. We show for the first time incorporation of GalNGc and GlcNGc into chondroitin/dermatan sulfates and heparan sulfates, respectively. As demonstrated by structural analysis, N-glycolylated hexosamines were found in cellular gangliosides and incorporated into Chinese hamster ovary cell O-glycans. Remarkably, GalNAc derivatives altered the overall O-glycosylation pattern as indicated by the occurrence of novel O-glycan structures. This study demonstrates that mammalian N-acetylhexosamine pathways and glycan assembly are surprisingly tolerant toward the N-glycolyl substituent.
引用
收藏
页码:28898 / 28916
页数:19
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