Glycobiology Introduction to GlycobiologyIntroduction to Glycobiology Glycan structuresGlycan structures Glycan functionsGlycan functions

Glycobiology Introduction to GlycobiologyIntroduction to Glycobiology Glycan structuresGlycan structures Glycan functionsGlycan functions

Light microscopy micrograph of Cryptococcus neoformans capsule delineated by India ink. The inner circle represents the fungal cell, with the wide outer circle being the capsule. Steenbergen et al. (2003) Microbes and Infection 5:667 Scanning electron microscopy of C. neoformans yeast cells. Van Duin et al. (2004) Antimicrobial Agents and Chemotherapy 48:2014 Light microscopy micrograph of Cryptococcus neoformans Steenbergen et al. (2003) Microbes and Infection 5:667 What if a MAJOR cell component was invisible by standard microscopy?

The Eukaryotic Cell Surface Tiny sugar “decorations”.. added as an afterthought Lodish, et al. (1995) Molecular Cell Biology 3rd ed. ?

The “glycocalyx” surrounding a fibroblast – glycans stained black lipid bilayer fibroblast extracellular intracellular Martinez-Palomo, A., et al. Cancer Res. 29, , 1969 The Cell Surface -- The Real Picture “Evolution has failed to generate a living cell devoid of surface glycosylation” - A. Varki “Every eukaryotic cell is covered with a dense and complex array of glycans, which also feature prominently in extracellular matrix and secreted soluble molecules.” … in fact…

Tropical Forest Canopy Ajit Varki, “Perspectives on the Future of Glycobiology,” Athens, GA, 14-Mar-2011

Tropical Forest Canopy Cohen & Varki (2010) OMICS 4:455

lipid bilayer fibroblastintracellular Martinez-Palomo, A., et al. Cancer Res. 29, , 1969 The Cell Surface Real image textbook image

Sharon and Lis (1993) Scientific American Cell-cell recognition is one important function Lectin Glycan

Life cycle of the influenza virus - the role of glycans

Influenza jumps from animals to humans via a slight change in glycan binding specificity Stevens, et al. (2006) Nat Rev Microbiol. 4:857-64

Influenza jumps from animals to humans via a slight change in glycan binding specificity Stevens, et al. (2006) Nat Rev Microbiol. 4:857-64

Inflammation: How we fight bacterial infection

Leukocyte cell surface glycan ligand required for endothelial recognition R

Marquardt, et al. (1999) J. Pediatr. 134, 681 You fight bacterial infection using glycans. This child could not …

Marquardt, et al. (1999) Blood 94, 3976 …until he was fed FUCOSE (a sugar), after which… “…neutrophil levels returned to the normal range.... there were no further infections”

CD59, a complement defense glycoprotein 20 kDa protein 3 kDa N-linked glycan 1 kDa O-linked glycan 1.5 kDa GPI anchor Proteins typically fold onto themselves, whereas glycans spread out in space plasma membrane

GlycoproteinGlycolipid

Glycobiology Introduction to GlycobiologyIntroduction to Glycobiology Glycan structuresGlycan structures Glycan functionsGlycan functions

Glycobiology Introduction to GlycobiologyIntroduction to Glycobiology Glycan structuresGlycan structures Glycan functionsGlycan functions

Monosaccharides equilibrate between open chain and ring forms. For each monosaccharide, two ring forms (anomeric configurations) are possible, α and β

Seven sugars and their structural relationship to glucose

Two important terminal sugars

When a glycosidic bond is formed, the anomeric configuration is “locked”

Nomenclature: -Name the non-reducing (left-most) sugar (Gal) -Name the anomeric configuration (β) -Name the anomeric carbon number (1) -Name the substituted carbon number (4) -Name the substituted sugar (Glc) -Name the non-reducing (left-most) sugar (Gal) -Name the anomeric configuration (β) -Name the anomeric carbon number (1) -Name the substituted carbon number (4) -Name the substituted sugar (Glc) RESULT: Gal β1-4 Glc

NeuAc α2-3 Gal β1-4 (Fuc α1-3) GlcNAc Nomenclature: Branches are placed in parentheses

NeuAc α2-3 Gal β1-4 (Fuc α1-3) GlcNAc

Molecular diversity glycans PolypeptidesGlycans Building blocks amino acids monosaccharides Number of different monomers 20 common 9 common Linkage sites per monomer 13-4 Possible linkage configurations 12 Possible homodimer structures 16-8 Linkage modes linear linear or branched

Three different amino acids (Ala, Ser, Tyr) – 6 structures Ala-Ser-TyrAla-Tyr-SerSer-Ala-TyrSer-Tyr-AlaTyr-Ala-SerTyr-Ser-Ala Three different sugars (Glc, Gal, Man) – 1,056 structures …

Gal a1-2 Glc a1-2 Man a1- Gal a1-3 Glc a1-2 Man a1- Gal a1-4 Glc a1-2 Man a1- Gal a1-6 Glc a1-2 Man a1- Gal b1-2 Glc a1-2 Man a1- Gal b1-3 Glc a1-2 Man a1- Gal b1-4 Glc a1-2 Man a1- Gal b1-6 Glc a1-2 Man a1- Gal a1-2 Glc a1-3 Man a1- Gal a1-3 Glc a1-3 Man a1- Gal a1-4 Glc a1-3 Man a1- Gal a1-6 Glc a1-3 Man a1- Gal b1-2 Glc a1-3 Man a1- Gal b1-3 Glc a1-3 Man a1- Gal b1-4 Glc a1-3 Man a1- Gal b1-6 Glc a1-3 Man a1- Gal a1-2 Glc a1-4 Man a1- Gal a1-3 Glc a1-4 Man a1- Gal a1-4 Glc a1-4 Man a1- Gal a1-6 Glc a1-4 Man a1- Gal b1-2 Glc a1-4 Man a1- Gal b1-3 Glc a1-4 Man a1- Gal b1-4 Glc a1-4 Man a1- Gal b1-6 Glc a1-4 Man a1- Gal a1-2 Glc a1-6 Man a1- Gal a1-3 Glc a1-6 Man a1- Gal a1-4 Glc a1-6 Man a1- Gal a1-6 Glc a1-6 Man a1- Gal b1-2 Glc a1-6 Man a1- Gal b1-3 Glc a1-6 Man a1- Gal b1-4 Glc a1-6 Man a1- Gal b1-6 Glc a1-6 Man a1- Gal a1-2 Glc b1-2 Man a1- Gal a1-3 Glc b1-2 Man a1- Gal a1-4 Glc b1-2 Man a1- Gal a1-6 Glc b1-2 Man a1- Gal b1-2 Glc b1-2 Man a1- Gal b1-3 Glc b1-2 Man a1- Gal b1-4 Glc b1-2 Man a1- Gal b1-6 Glc b1-2 Man a1- Gal a1-2 Glc b1-3 Man a1- Gal a1-3 Glc b1-3 Man a1- Gal a1-4 Glc b1-3 Man a1- Gal a1-6 Glc b1-3 Man a1- Gal b1-2 Glc b1-3 Man a1- Gal b1-3 Glc b1-3 Man a1- Gal b1-4 Glc b1-3 Man a1- Gal b1-6 Glc b1-3 Man a1- Gal a1-2 Glc b1-4 Man a1- Gal a1-3 Glc b1-4 Man a1- Gal a1-4 Glc b1-4 Man a1- Gal a1-6 Glc b1-4 Man a1- Gal b1-2 Glc b1-4 Man a1- Gal b1-3 Glc b1-4 Man a1- Gal b1-4 Glc b1-4 Man a1- Gal b1-6 Glc b1-4 Man a1- Gal a1-2 Glc b1-6 Man a1- Gal a1-3 Glc b1-6 Man a1- Gal a1-4 Glc b1-6 Man a1- Gal a1-6 Glc b1-6 Man a1- Gal b1-2 Glc b1-6 Man a1- Gal b1-3 Glc b1-6 Man a1- Gal b1-4 Glc b1-6 Man a1- Gal b1-6 Glc b1-6 Man a1- Gal a1-2 Glc a1-2 Man b1- Gal a1-3 Glc a1-2 Man b1- Gal a1-4 Glc a1-2 Man b1- Gal a1-6 Glc a1-2 Man b1- Gal b1-2 Glc a1-2 Man b1- Gal b1-3 Glc a1-2 Man b1- Gal b1-4 Glc a1-2 Man b1- Gal b1-6 Glc a1-2 Man b1- Gal a1-2 Glc a1-3 Man b1- Gal a1-3 Glc a1-3 Man b1- Gal a1-4 Glc a1-3 Man b1- Gal a1-6 Glc a1-3 Man b1- Gal b1-2 Glc a1-3 Man b1- Gal b1-3 Glc a1-3 Man b1- Gal b1-4 Glc a1-3 Man b1- Gal b1-6 Glc a1-3 Man b1- Gal a1-2 Glc a1-4 Man b1- Gal a1-3 Glc a1-4 Man b1- Gal a1-4 Glc a1-4 Man b1- Gal a1-6 Glc a1-4 Man b1- Gal b1-2 Glc a1-4 Man b1- Gal b1-3 Glc a1-4 Man b1- Gal b1-4 Glc a1-4 Man b1- Gal b1-6 Glc a1-4 Man b1- Gal a1-2 Glc a1-6 Man b1- Gal a1-3 Glc a1-6 Man b1- Gal a1-4 Glc a1-6 Man b1- Gal a1-6 Glc a1-6 Man b1- Gal b1-2 Glc a1-6 Man b1- Gal b1-3 Glc a1-6 Man b1- Gal b1-4 Glc a1-6 Man b1- Gal b1-6 Glc a1-6 Man b1- Gal a1-2 Glc b1-2 Man b1- Gal a1-3 Glc b1-2 Man b1- Gal a1-4 Glc b1-2 Man b1- Gal a1-6 Glc b1-2 Man b1- Gal b1-2 Glc b1-2 Man b1- Gal b1-3 Glc b1-2 Man b1- Gal b1-4 Glc b1-2 Man b1- Gal b1-6 Glc b1-2 Man b1- Gal a1-2 Glc b1-3 Man b1- Gal a1-3 Glc b1-3 Man b1- Gal a1-4 Glc b1-3 Man b1- Gal a1-6 Glc b1-3 Man b1- Gal b1-2 Glc b1-3 Man b1- Gal b1-3 Glc b1-3 Man b1- Gal b1-4 Glc b1-3 Man b1- Gal b1-6 Glc b1-3 Man b1- Gal a1-2 Glc b1-4 Man b1- Gal a1-3 Glc b1-4 Man b1- Gal a1-4 Glc b1-4 Man b1- Gal a1-6 Glc b1-4 Man b1- Gal b1-2 Glc b1-4 Man b1- Gal b1-3 Glc b1-4 Man b1- Gal b1-4 Glc b1-4 Man b1- Gal b1-6 Glc b1-4 Man b1- Gal a1-2 Glc b1-6 Man b1- Gal a1-3 Glc b1-6 Man b1- Gal a1-4 Glc b1-6 Man b1- Gal a1-6 Glc b1-6 Man b1- Gal b1-2 Glc b1-6 Man b1- Gal b1-3 Glc b1-6 Man b1- Gal b1-4 Glc b1-6 Man b1- Gal b1-6 Glc b1-6 Man b1- Gal a1-2 Man a1-2 Glc a1- Gal a1-3 Man a1-2 Glc a1- Gal a1-4 Man a1-2 Glc a1- Gal a1-6 Man a1-2 Glc a1- Gal b1-2 Man a1-2 Glc a1- Gal b1-3 Man a1-2 Glc a1- Gal b1-4 Man a1-2 Glc a1- Gal b1-6 Man a1-2 Glc a1- Gal a1-2 Man a1-3 Glc a1- Gal a1-3 Man a1-3 Glc a1- Gal a1-4 Man a1-3 Glc a1- Gal a1-6 Man a1-3 Glc a1- Gal b1-2 Man a1-3 Glc a1- Gal b1-3 Man a1-3 Glc a1- Gal b1-4 Man a1-3 Glc a1- Gal b1-6 Man a1-3 Glc a1- Gal a1-2 Man a1-4 Glc a1- Gal a1-3 Man a1-4 Glc a1- Gal a1-4 Man a1-4 Glc a1- Gal a1-6 Man a1-4 Glc a1- Gal b1-2 Man a1-4 Glc a1- Gal b1-3 Man a1-4 Glc a1- Gal b1-4 Man a1-4 Glc a1- Gal b1-6 Man a1-4 Glc a1- Gal a1-2 Man a1-6 Glc a1- Gal a1-3 Man a1-6 Glc a1- Gal a1-4 Man a1-6 Glc a1- Gal a1-6 Man a1-6 Glc a1- Gal b1-2 Man a1-6 Glc a1- Gal b1-3 Man a1-6 Glc a1- Gal b1-4 Man a1-6 Glc a1- Gal b1-6 Man a1-6 Glc a1- Gal a1-2 Man b1-2 Glc a1- Gal a1-3 Man b1-2 Glc a1- Gal a1-4 Man b1-2 Glc a1- Gal a1-6 Man b1-2 Glc a1- Gal b1-2 Man b1-2 Glc a1- Gal b1-3 Man b1-2 Glc a1- Gal b1-4 Man b1-2 Glc a1- Gal b1-6 Man b1-2 Glc a1- Gal a1-2 Man b1-3 Glc a1- Gal a1-3 Man b1-3 Glc a1- Gal a1-4 Man b1-3 Glc a1- Gal a1-6 Man b1-3 Glc a1- Gal b1-2 Man b1-3 Glc a1- Gal b1-3 Man b1-3 Glc a1- Gal b1-4 Man b1-3 Glc a1- Gal b1-6 Man b1-3 Glc a1- Gal a1-2 Man b1-4 Glc a1- Gal a1-3 Man b1-4 Glc a1- Gal a1-4 Man b1-4 Glc a1- Gal a1-6 Man b1-4 Glc a1- Gal b1-2 Man b1-4 Glc a1- Gal b1-3 Man b1-4 Glc a1- Gal b1-4 Man b1-4 Glc a1- Gal b1-6 Man b1-4 Glc a1- Gal a1-2 Man b1-6 Glc a1- Gal a1-3 Man b1-6 Glc a1- Gal a1-4 Man b1-6 Glc a1- Gal a1-6 Man b1-6 Glc a1- Gal b1-2 Man b1-6 Glc a1- Gal b1-3 Man b1-6 Glc a1- Gal b1-4 Man b1-6 Glc a1- Gal b1-6 Man b1-6 Glc a1- Gal a1-2 Man a1-2 Glc b1- Gal a1-3 Man a1-2 Glc b1- Gal a1-4 Man a1-2 Glc b1- Gal a1-6 Man a1-2 Glc b1- Gal b1-2 Man a1-2 Glc b1- Gal b1-3 Man a1-2 Glc b1- Gal b1-4 Man a1-2 Glc b1- Gal b1-6 Man a1-2 Glc b1- Gal a1-2 Man a1-3 Glc b1- Gal a1-3 Man a1-3 Glc b1- Gal a1-4 Man a1-3 Glc b1- Gal a1-6 Man a1-3 Glc b1- Gal b1-2 Man a1-3 Glc b1- Gal b1-3 Man a1-3 Glc b1- Gal b1-4 Man a1-3 Glc b1- Gal b1-6 Man a1-3 Glc b1- Gal a1-2 Man a1-4 Glc b1- Gal a1-3 Man a1-4 Glc b1- Gal a1-4 Man a1-4 Glc b1- Gal a1-6 Man a1-4 Glc b1- Gal b1-2 Man a1-4 Glc b1- Gal b1-3 Man a1-4 Glc b1- Gal b1-4 Man a1-4 Glc b1- Gal b1-6 Man a1-4 Glc b1- Gal a1-2 Man a1-6 Glc b1- Gal a1-3 Man a1-6 Glc b1- Gal a1-4 Man a1-6 Glc b1- Gal a1-6 Man a1-6 Glc b1- Gal b1-2 Man a1-6 Glc b1- Gal b1-3 Man a1-6 Glc b1- Gal b1-4 Man a1-6 Glc b1- Gal b1-6 Man a1-6 Glc b1- Gal a1-2 Man b1-2 Glc b1- Gal a1-3 Man b1-2 Glc b1- Gal a1-4 Man b1-2 Glc b1- Gal a1-6 Man b1-2 Glc b1- Gal b1-2 Man b1-2 Glc b1- Gal b1-3 Man b1-2 Glc b1- Gal b1-4 Man b1-2 Glc b1- Gal b1-6 Man b1-2 Glc b1- Gal a1-2 Man b1-3 Glc b1- Gal a1-3 Man b1-3 Glc b1- Gal a1-4 Man b1-3 Glc b1- Gal a1-6 Man b1-3 Glc b1- Gal b1-2 Man b1-3 Glc b1- Gal b1-3 Man b1-3 Glc b1- Gal b1-4 Man b1-3 Glc b1- Gal b1-6 Man b1-3 Glc b1- Gal a1-2 Man b1-4 Glc b1- Gal a1-3 Man b1-4 Glc b1- Gal a1-4 Man b1-4 Glc b1- Gal a1-6 Man b1-4 Glc b1- Gal b1-2 Man b1-4 Glc b1- Gal b1-3 Man b1-4 Glc b1- Gal b1-4 Man b1-4 Glc b1- Gal b1-6 Man b1-4 Glc b1- Gal a1-2 Man b1-6 Glc b1- Gal a1-3 Man b1-6 Glc b1- Gal a1-4 Man b1-6 Glc b1- Gal a1-6 Man b1-6 Glc b1- Gal b1-2 Man b1-6 Glc b1- Gal b1-3 Man b1-6 Glc b1- Gal b1-4 Man b1-6 Glc b1- Gal b1-6 Man b1-6 Glc b1- Man a1-2 Glc a1-2 Gal a1- Man a1-3 Glc a1-2 Gal a1- Man a1-4 Glc a1-2 Gal a1- Man a1-6 Glc a1-2 Gal a1- Man b1-2 Glc a1-2 Gal a1- Man b1-3 Glc a1-2 Gal a1- Man b1-4 Glc a1-2 Gal a1- Man b1-6 Glc a1-2 Gal a1- Man a1-2 Glc a1-3 Gal a1- Man a1-3 Glc a1-3 Gal a1- Man a1-4 Glc a1-3 Gal a1- Man a1-6 Glc a1-3 Gal a1- Man b1-2 Glc a1-3 Gal a1- Man b1-3 Glc a1-3 Gal a1- Man b1-4 Glc a1-3 Gal a1- Man b1-6 Glc a1-3 Gal a1- Man a1-2 Glc a1-4 Gal a1- Man a1-3 Glc a1-4 Gal a1- Man a1-4 Glc a1-4 Gal a1- Man a1-6 Glc a1-4 Gal a1- Man b1-2 Glc a1-4 Gal a1- Man b1-3 Glc a1-4 Gal a1- Man b1-4 Glc a1-4 Gal a1- Man b1-6 Glc a1-4 Gal a1- Man a1-2 Glc a1-6 Gal a1- Man a1-3 Glc a1-6 Gal a1- Man a1-4 Glc a1-6 Gal a1- Man a1-6 Glc a1-6 Gal a1- Man b1-2 Glc a1-6 Gal a1- Man b1-3 Glc a1-6 Gal a1- Man b1-4 Glc a1-6 Gal a1- Man b1-6 Glc a1-6 Gal a1- Man a1-2 Glc b1-2 Gal a1- Man a1-3 Glc b1-2 Gal a1- Man a1-4 Glc b1-2 Gal a1- Man a1-6 Glc b1-2 Gal a1- Man b1-2 Glc b1-2 Gal a1- Man b1-3 Glc b1-2 Gal a1- Man b1-4 Glc b1-2 Gal a1- Man b1-6 Glc b1-2 Gal a1- Man a1-2 Glc b1-3 Gal a1- Man a1-3 Glc b1-3 Gal a1- Man a1-4 Glc b1-3 Gal a1- Man a1-6 Glc b1-3 Gal a1- Man b1-2 Glc b1-3 Gal a1- Man b1-3 Glc b1-3 Gal a1- Man b1-4 Glc b1-3 Gal a1- Man b1-6 Glc b1-3 Gal a1- Man a1-2 Glc b1-4 Gal a1- Man a1-3 Glc b1-4 Gal a1- Man a1-4 Glc b1-4 Gal a1- Man a1-6 Glc b1-4 Gal a1- Man b1-2 Glc b1-4 Gal a1- Man b1-3 Glc b1-4 Gal a1- Man b1-4 Glc b1-4 Gal a1- Man b1-6 Glc b1-4 Gal a1- Man a1-2 Glc b1-6 Gal a1- Man a1-3 Glc b1-6 Gal a1- Man a1-4 Glc b1-6 Gal a1- Man a1-6 Glc b1-6 Gal a1- Man b1-2 Glc b1-6 Gal a1- Man b1-3 Glc b1-6 Gal a1- Man b1-4 Glc b1-6 Gal a1- Man b1-6 Glc b1-6 Gal a1- Man a1-2 Glc a1-2 Gal b1- Man a1-3 Glc a1-2 Gal b1- Man a1-4 Glc a1-2 Gal b1- Man a1-6 Glc a1-2 Gal b1- Man b1-2 Glc a1-2 Gal b1- Man b1-3 Glc a1-2 Gal b1- Man b1-4 Glc a1-2 Gal b1- Man b1-6 Glc a1-2 Gal b1- Man a1-2 Glc a1-3 Gal b1- Man a1-3 Glc a1-3 Gal b1- Man a1-4 Glc a1-3 Gal b1- Man a1-6 Glc a1-3 Gal b1- Man b1-2 Glc a1-3 Gal b1- Man b1-3 Glc a1-3 Gal b1- Man b1-4 Glc a1-3 Gal b1- Man b1-6 Glc a1-3 Gal b1- Man a1-2 Glc a1-4 Gal b1- Man a1-3 Glc a1-4 Gal b1- Man a1-4 Glc a1-4 Gal b1- Man a1-6 Glc a1-4 Gal b1- Man b1-2 Glc a1-4 Gal b1- Man b1-3 Glc a1-4 Gal b1- Man b1-4 Glc a1-4 Gal b1- Man b1-6 Glc a1-4 Gal b1- Man a1-2 Glc a1-6 Gal b1- Man a1-3 Glc a1-6 Gal b1- Man a1-4 Glc a1-6 Gal b1- Man a1-6 Glc a1-6 Gal b1- Man b1-2 Glc a1-6 Gal b1- Man b1-3 Glc a1-6 Gal b1- Man b1-4 Glc a1-6 Gal b1- Man b1-6 Glc a1-6 Gal b1- Man a1-2 Glc b1-2 Gal b1- Man a1-3 Glc b1-2 Gal b1- Man a1-4 Glc b1-2 Gal b1- Man a1-6 Glc b1-2 Gal b1- Man b1-2 Glc b1-2 Gal b1- Man b1-3 Glc b1-2 Gal b1- Man b1-4 Glc b1-2 Gal b1- Man b1-6 Glc b1-2 Gal b1- Man a1-2 Glc b1-3 Gal b1- Man a1-3 Glc b1-3 Gal b1- Man a1-4 Glc b1-3 Gal b1- Man a1-6 Glc b1-3 Gal b1- Man b1-2 Glc b1-3 Gal b1- Man b1-3 Glc b1-3 Gal b1- Man b1-4 Glc b1-3 Gal b1- Man b1-6 Glc b1-3 Gal b1- Man a1-2 Glc b1-4 Gal b1- Man a1-3 Glc b1-4 Gal b1- Man a1-4 Glc b1-4 Gal b1- Man a1-6 Glc b1-4 Gal b1- Man b1-2 Glc b1-4 Gal b1- Man b1-3 Glc b1-4 Gal b1- Man b1-4 Glc b1-4 Gal b1- Man b1-6 Glc b1-4 Gal b1- Man a1-2 Glc b1-6 Gal b1- Man a1-3 Glc b1-6 Gal b1- Man a1-4 Glc b1-6 Gal b1- Man a1-6 Glc b1-6 Gal b1- Man b1-2 Glc b1-6 Gal b1- Man b1-3 Glc b1-6 Gal b1- Man b1-4 Glc b1-6 Gal b1- Man b1-6 Glc b1-6 Gal b1- Man a1-2 Gal a1-2 Glc a1- Man a1-3 Gal a1-2 Glc a1- Man a1-4 Gal a1-2 Glc a1- Man a1-6 Gal a1-2 Glc a1- Man b1-2 Gal a1-2 Glc a1- Man b1-3 Gal a1-2 Glc a1- Man b1-4 Gal a1-2 Glc a1- Man b1-6 Gal a1-2 Glc a1- Man a1-2 Gal a1-3 Glc a1- Man a1-3 Gal a1-3 Glc a1- Man a1-4 Gal a1-3 Glc a1- Man a1-6 Gal a1-3 Glc a1- Man b1-2 Gal a1-3 Glc a1- Man b1-3 Gal a1-3 Glc a1- Man b1-4 Gal a1-3 Glc a1- Man b1-6 Gal a1-3 Glc a1- Man a1-2 Gal a1-4 Glc a1- Man a1-3 Gal a1-4 Glc a1- Man a1-4 Gal a1-4 Glc a1- Man a1-6 Gal a1-4 Glc a1- Man b1-2 Gal a1-4 Glc a1- Man b1-3 Gal a1-4 Glc a1- Man b1-4 Gal a1-4 Glc a1- Man b1-6 Gal a1-4 Glc a1- Man a1-2 Gal a1-6 Glc a1- Man a1-3 Gal a1-6 Glc a1- Man a1-4 Gal a1-6 Glc a1- Man a1-6 Gal a1-6 Glc a1- Man b1-2 Gal a1-6 Glc a1- Man b1-3 Gal a1-6 Glc a1- Man b1-4 Gal a1-6 Glc a1- Man b1-6 Gal a1-6 Glc a1- Man a1-2 Gal b1-2 Glc a1- Man a1-3 Gal b1-2 Glc a1- Man a1-4 Gal b1-2 Glc a1- Man a1-6 Gal b1-2 Glc a1- Man b1-2 Gal b1-2 Glc a1- Man b1-3 Gal b1-2 Glc a1- Man b1-4 Gal b1-2 Glc a1- Man b1-6 Gal b1-2 Glc a1- Man a1-2 Gal b1-3 Glc a1- Man a1-3 Gal b1-3 Glc a1- Man a1-4 Gal b1-3 Glc a1- Man a1-6 Gal b1-3 Glc a1- Man b1-2 Gal b1-3 Glc a1- Man b1-3 Gal b1-3 Glc a1- Man b1-4 Gal b1-3 Glc a1- Man b1-6 Gal b1-3 Glc a1- Man a1-2 Gal b1-4 Glc a1- Man a1-3 Gal b1-4 Glc a1- Man a1-4 Gal b1-4 Glc a1- Man a1-6 Gal b1-4 Glc a1- Man b1-2 Gal b1-4 Glc a1- Man b1-3 Gal b1-4 Glc a1- Man b1-4 Gal b1-4 Glc a1- Man b1-6 Gal b1-4 Glc a1- Man a1-2 Gal b1-6 Glc a1- Man a1-3 Gal b1-6 Glc a1- Man a1-4 Gal b1-6 Glc a1- Man a1-6 Gal b1-6 Glc a1- Man b1-2 Gal b1-6 Glc a1- Man b1-3 Gal b1-6 Glc a1- Man b1-4 Gal b1-6 Glc a1- Man b1-6 Gal b1-6 Glc a1- Man a1-2 Gal a1-2 Glc b1 Man a1-3 Gal a1-2 Glc b1 Man a1-4 Gal a1-2 Glc b1 Man a1-6 Gal a1-2 Glc b1 Man b1-2 Gal a1-2 Glc b1 Man b1-3 Gal a1-2 Glc b1 Man b1-4 Gal a1-2 Glc b1 Man b1-6 Gal a1-2 Glc b1 Man a1-2 Gal a1-3 Glc b1 Man a1-3 Gal a1-3 Glc b1 Man a1-4 Gal a1-3 Glc b1 Man a1-6 Gal a1-3 Glc b1 Man b1-2 Gal a1-3 Glc b1 Man b1-3 Gal a1-3 Glc b1 Man b1-4 Gal a1-3 Glc b1 Man b1-6 Gal a1-3 Glc b1 Man a1-2 Gal a1-4 Glc b1 Man a1-3 Gal a1-4 Glc b1 Man a1-4 Gal a1-4 Glc b1 Man a1-6 Gal a1-4 Glc b1 Man b1-2 Gal a1-4 Glc b1 Man b1-3 Gal a1-4 Glc b1 Man b1-4 Gal a1-4 Glc b1 Man b1-6 Gal a1-4 Glc b1 Man a1-2 Gal a1-6 Glc b1 Man a1-3 Gal a1-6 Glc b1 Man a1-4 Gal a1-6 Glc b1 Man a1-6 Gal a1-6 Glc b1 Man b1-2 Gal a1-6 Glc b1 Man b1-3 Gal a1-6 Glc b1 Man b1-4 Gal a1-6 Glc b1 Man b1-6 Gal a1-6 Glc b1 Man a1-2 Gal b1-2 Glc b1 Man a1-3 Gal b1-2 Glc b1 Man a1-4 Gal b1-2 Glc b1 Man a1-6 Gal b1-2 Glc b1 Man b1-2 Gal b1-2 Glc b1 Man b1-3 Gal b1-2 Glc b1 Man b1-4 Gal b1-2 Glc b1 Man b1-6 Gal b1-2 Glc b1 Man a1-2 Gal b1-3 Glc b1 Man a1-3 Gal b1-3 Glc b1 Man a1-4 Gal b1-3 Glc b1 Man a1-6 Gal b1-3 Glc b1 Man b1-2 Gal b1-3 Glc b1 Man b1-3 Gal b1-3 Glc b1 Man b1-4 Gal b1-3 Glc b1 Man b1-6 Gal b1-3 Glc b1 Man a1-2 Gal b1-4 Glc b1 Man a1-3 Gal b1-4 Glc b1 Man a1-4 Gal b1-4 Glc b1 Man a1-6 Gal b1-4 Glc b1 Man b1-2 Gal b1-4 Glc b1 Man b1-3 Gal b1-4 Glc b1 Man b1-4 Gal b1-4 Glc b1 Man b1-6 Gal b1-4 Glc b1 Man a1-2 Gal b1-6 Glc b1 Man a1-3 Gal b1-6 Glc b1 Man a1-4 Gal b1-6 Glc b1 Man a1-6 Gal b1-6 Glc b1 Man b1-2 Gal b1-6 Glc b1 Man b1-3 Gal b1-6 Glc b1 Man b1-4 Gal b1-6 Glc b1 Man b1-6 Gal b1-6 Glc b1 Glc a1-2 Man a1-2 Gal a1- Glc a1-3 Man a1-2 Gal a1- Glc a1-4 Man a1-2 Gal a1- Glc a1-6 Man a1-2 Gal a1- Glc b1-2 Man a1-2 Gal a1- Glc b1-3 Man a1-2 Gal a1- Glc b1-4 Man a1-2 Gal a1- Glc b1-6 Man a1-2 Gal a1- Glc a1-2 Man a1-3 Gal a1- Glc a1-3 Man a1-3 Gal a1- Glc a1-4 Man a1-3 Gal a1- Glc a1-6 Man a1-3 Gal a1- Glc b1-2 Man a1-3 Gal a1- Glc b1-3 Man a1-3 Gal a1- Glc b1-4 Man a1-3 Gal a1- Glc b1-6 Man a1-3 Gal a1- Glc a1-2 Man a1-4 Gal a1- Glc a1-3 Man a1-4 Gal a1- Glc a1-4 Man a1-4 Gal a1- Glc a1-6 Man a1-4 Gal a1- Glc b1-2 Man a1-4 Gal a1- Glc b1-3 Man a1-4 Gal a1- Glc b1-4 Man a1-4 Gal a1- Glc b1-6 Man a1-4 Gal a1- Glc a1-2 Man a1-6 Gal a1- Glc a1-3 Man a1-6 Gal a1- Glc a1-4 Man a1-6 Gal a1- Glc a1-6 Man a1-6 Gal a1- Glc b1-2 Man a1-6 Gal a1- Glc b1-3 Man a1-6 Gal a1- Glc b1-4 Man a1-6 Gal a1- Glc b1-6 Man a1-6 Gal a1- Glc a1-2 Man b1-2 Gal a1- Glc a1-3 Man b1-2 Gal a1- Glc a1-4 Man b1-2 Gal a1- Glc a1-6 Man b1-2 Gal a1- Glc b1-2 Man b1-2 Gal a1- Glc b1-3 Man b1-2 Gal a1- Glc b1-4 Man b1-2 Gal a1- Glc b1-6 Man b1-2 Gal a1- Glc a1-2 Man b1-3 Gal a1- Glc a1-3 Man b1-3 Gal a1- Glc a1-4 Man b1-3 Gal a1- Glc a1-6 Man b1-3 Gal a1- Glc b1-2 Man b1-3 Gal a1- Glc b1-3 Man b1-3 Gal a1- Glc b1-4 Man b1-3 Gal a1- Glc b1-6 Man b1-3 Gal a1- Glc a1-2 Man b1-4 Gal a1- Glc a1-3 Man b1-4 Gal a1- Glc a1-4 Man b1-4 Gal a1- Glc a1-6 Man b1-4 Gal a1- Glc b1-2 Man b1-4 Gal a1- Glc b1-3 Man b1-4 Gal a1- Glc b1-4 Man b1-4 Gal a1- Glc b1-6 Man b1-4 Gal a1- Glc a1-2 Man b1-6 Gal a1- Glc a1-3 Man b1-6 Gal a1- Glc a1-4 Man b1-6 Gal a1- Glc a1-6 Man b1-6 Gal a1- Glc b1-2 Man b1-6 Gal a1- Glc b1-3 Man b1-6 Gal a1- Glc b1-4 Man b1-6 Gal a1- Glc b1-6 Man b1-6 Gal a1- Glc a1-2 Man a1-2 Gal b1- Glc a1-3 Man a1-2 Gal b1- Glc a1-4 Man a1-2 Gal b1- Glc a1-6 Man a1-2 Gal b1- Glc b1-2 Man a1-2 Gal b1- Glc b1-3 Man a1-2 Gal b1- Glc b1-4 Man a1-2 Gal b1- Glc b1-6 Man a1-2 Gal b1- Glc a1-2 Man a1-3 Gal b1- Glc a1-3 Man a1-3 Gal b1- Glc a1-4 Man a1-3 Gal b1- Glc a1-6 Man a1-3 Gal b1- Glc b1-2 Man a1-3 Gal b1- Glc b1-3 Man a1-3 Gal b1- Glc b1-4 Man a1-3 Gal b1- Glc b1-6 Man a1-3 Gal b1- Glc a1-2 Man a1-4 Gal b1- Glc a1-3 Man a1-4 Gal b1- Glc a1-4 Man a1-4 Gal b1- Glc a1-6 Man a1-4 Gal b1- Glc b1-2 Man a1-4 Gal b1- Glc b1-3 Man a1-4 Gal b1- Glc b1-4 Man a1-4 Gal b1- Glc b1-6 Man a1-4 Gal b1- Glc a1-2 Man a1-6 Gal b1- Glc a1-3 Man a1-6 Gal b1- Glc a1-4 Man a1-6 Gal b1- Glc a1-6 Man a1-6 Gal b1- Glc b1-2 Man a1-6 Gal b1- Glc b1-3 Man a1-6 Gal b1- Glc b1-4 Man a1-6 Gal b1- Glc b1-6 Man a1-6 Gal b1- Glc a1-2 Man b1-2 Gal b1- Glc a1-3 Man b1-2 Gal b1- Glc a1-4 Man b1-2 Gal b1- Glc a1-6 Man b1-2 Gal b1- Glc b1-2 Man b1-2 Gal b1- Glc b1-3 Man b1-2 Gal b1- Glc b1-4 Man b1-2 Gal b1- Glc b1-6 Man b1-2 Gal b1- Glc a1-2 Man b1-3 Gal b1- Glc a1-3 Man b1-3 Gal b1- Glc a1-4 Man b1-3 Gal b1- Glc a1-6 Man b1-3 Gal b1- Glc b1-2 Man b1-3 Gal b1- Glc b1-3 Man b1-3 Gal b1- Glc b1-4 Man b1-3 Gal b1- Glc b1-6 Man b1-3 Gal b1- Glc a1-2 Man b1-4 Gal b1- Glc a1-3 Man b1-4 Gal b1- Glc a1-4 Man b1-4 Gal b1- Glc a1-6 Man b1-4 Gal b1- Glc b1-2 Man b1-4 Gal b1- Glc b1-3 Man b1-4 Gal b1- Glc b1-4 Man b1-4 Gal b1- Glc b1-6 Man b1-4 Gal b1- Glc a1-2 Man b1-6 Gal b1- Glc a1-3 Man b1-6 Gal b1- Glc a1-4 Man b1-6 Gal b1- Glc a1-6 Man b1-6 Gal b1- Glc b1-2 Man b1-6 Gal b1- Glc b1-3 Man b1-6 Gal b1- Glc b1-4 Man b1-6 Gal b1- Glc b1-6 Man b1-6 Gal b1- Glc a1-2 Gal a1-2 Man a1- Glc a1-3 Gal a1-2 Man a1- Glc a1-4 Gal a1-2 Man a1- Glc a1-6 Gal a1-2 Man a1- Glc b1-2 Gal a1-2 Man a1- Glc b1-3 Gal a1-2 Man a1- Glc b1-4 Gal a1-2 Man a1- Glc b1-6 Gal a1-2 Man a1- Glc a1-2 Gal a1-3 Man a1- Glc a1-3 Gal a1-3 Man a1- Glc a1-4 Gal a1-3 Man a1- Glc a1-6 Gal a1-3 Man a1- Glc b1-2 Gal a1-3 Man a1- Glc b1-3 Gal a1-3 Man a1- Glc b1-4 Gal a1-3 Man a1- Glc b1-6 Gal a1-3 Man a1- Glc a1-2 Gal a1-4 Man a1- Glc a1-3 Gal a1-4 Man a1- Glc a1-4 Gal a1-4 Man a1- Glc a1-6 Gal a1-4 Man a1- Glc b1-2 Gal a1-4 Man a1- Glc b1-3 Gal a1-4 Man a1- Glc b1-4 Gal a1-4 Man a1- Glc b1-6 Gal a1-4 Man a1- Glc a1-2 Gal a1-6 Man a1- Glc a1-3 Gal a1-6 Man a1- Glc a1-4 Gal a1-6 Man a1- Glc a1-6 Gal a1-6 Man a1- Glc b1-2 Gal a1-6 Man a1- Glc b1-3 Gal a1-6 Man a1- Glc b1-4 Gal a1-6 Man a1- Glc b1-6 Gal a1-6 Man a1- Glc a1-2 Gal b1-2 Man a1- Glc a1-3 Gal b1-2 Man a1- Glc a1-4 Gal b1-2 Man a1- Glc a1-6 Gal b1-2 Man a1- Glc b1-2 Gal b1-2 Man a1- Glc b1-3 Gal b1-2 Man a1- Glc b1-4 Gal b1-2 Man a1- Glc b1-6 Gal b1-2 Man a1- Glc a1-2 Gal b1-3 Man a1- Glc a1-3 Gal b1-3 Man a1- Glc a1-4 Gal b1-3 Man a1- Glc a1-6 Gal b1-3 Man a1- Glc b1-2 Gal b1-3 Man a1- Glc b1-3 Gal b1-3 Man a1- Glc b1-4 Gal b1-3 Man a1- Glc b1-6 Gal b1-3 Man a1- Glc a1-2 Gal b1-4 Man a1- Glc a1-3 Gal b1-4 Man a1- Glc a1-4 Gal b1-4 Man a1- Glc a1-6 Gal b1-4 Man a1- Glc b1-2 Gal b1-4 Man a1- Glc b1-3 Gal b1-4 Man a1- Glc b1-4 Gal b1-4 Man a1- Glc b1-6 Gal b1-4 Man a1- Glc a1-2 Gal b1-6 Man a1- Glc a1-3 Gal b1-6 Man a1- Glc a1-4 Gal b1-6 Man a1- Glc a1-6 Gal b1-6 Man a1- Glc b1-2 Gal b1-6 Man a1- Glc b1-3 Gal b1-6 Man a1- Glc b1-4 Gal b1-6 Man a1- Glc b1-6 Gal b1-6 Man a1- Glc a1-2 Gal a1-2 Man b1- Glc a1-3 Gal a1-2 Man b1- Glc a1-4 Gal a1-2 Man b1- Glc a1-6 Gal a1-2 Man b1- Glc b1-2 Gal a1-2 Man b1- Glc b1-3 Gal a1-2 Man b1- Glc b1-4 Gal a1-2 Man b1- Glc b1-6 Gal a1-2 Man b1- Glc a1-2 Gal a1-3 Man b1- Glc a1-3 Gal a1-3 Man b1- Glc a1-4 Gal a1-3 Man b1- Glc a1-6 Gal a1-3 Man b1- Glc b1-2 Gal a1-3 Man b1- Glc b1-3 Gal a1-3 Man b1- Glc b1-4 Gal a1-3 Man b1- Glc b1-6 Gal a1-3 Man b1- Glc a1-2 Gal a1-4 Man b1- Glc a1-3 Gal a1-4 Man b1- Glc a1-4 Gal a1-4 Man b1- Glc a1-6 Gal a1-4 Man b1- Glc b1-2 Gal a1-4 Man b1- Glc b1-3 Gal a1-4 Man b1- Glc b1-4 Gal a1-4 Man b1- Glc b1-6 Gal a1-4 Man b1- Glc a1-2 Gal a1-6 Man b1- Glc a1-3 Gal a1-6 Man b1- Glc a1-4 Gal a1-6 Man b1- Glc a1-6 Gal a1-6 Man b1- Glc b1-2 Gal a1-6 Man b1- Glc b1-3 Gal a1-6 Man b1- Glc b1-4 Gal a1-6 Man b1- Glc b1-6 Gal a1-6 Man b1- Glc a1-2 Gal b1-2 Man b1- Glc a1-3 Gal b1-2 Man b1- Glc a1-4 Gal b1-2 Man b1- Glc a1-6 Gal b1-2 Man b1- Glc b1-2 Gal b1-2 Man b1- Glc b1-3 Gal b1-2 Man b1- Glc b1-4 Gal b1-2 Man b1- Glc b1-6 Gal b1-2 Man b1- Glc a1-2 Gal b1-3 Man b1- Glc a1-3 Gal b1-3 Man b1- Glc a1-4 Gal b1-3 Man b1- Glc a1-6 Gal b1-3 Man b1- Glc b1-2 Gal b1-3 Man b1- Glc b1-3 Gal b1-3 Man b1- Glc b1-4 Gal b1-3 Man b1- Glc b1-6 Gal b1-3 Man b1- Glc a1-2 Gal b1-4 Man b1- Glc a1-3 Gal b1-4 Man b1- Glc a1-4 Gal b1-4 Man b1- Glc a1-6 Gal b1-4 Man b1- Glc b1-2 Gal b1-4 Man b1- Glc b1-3 Gal b1-4 Man b1- Glc b1-4 Gal b1-4 Man b1- Glc b1-6 Gal b1-4 Man b1- Glc a1-2 Gal b1-6 Man b1- Glc a1-3 Gal b1-6 Man b1- Glc a1-4 Gal b1-6 Man b1- Glc a1-6 Gal b1-6 Man b1- Glc b1-2 Gal b1-6 Man b1- Glc b1-3 Gal b1-6 Man b1- Glc b1-4 Gal b1-6 Man b1- Glc b1-6 Gal b1-6 Man b1- Gal a1-2 (Glc a1-3) Man a1- Gal a1-2 (Glc a1-4) Man a1- Gal a1-2 (Glc a1-6) Man a1- Gal a1-3 (Glc a1-2) Man a1- Gal a1-3 (Glc a1-4) Man a1- Gal a1-3 (Glc a1-6) Man a1- Gal a1-4 (Glc a1-2) Man a1- Gal a1-4 (Glc a1-3) Man a1- Gal a1-4 (Glc a1-6) Man a1- Gal a1-6 (Glc a1-2) Man a1- Gal a1-6 (Glc a1-3) Man a1- Gal a1-6 (Glc a1-4) Man a1- Gal b1-2 (Glc a1-3) Man a1- Gal b1-2 (Glc a1-4) Man a1- Gal b1-2 (Glc a1-6) Man a1- Gal b1-3 (Glc a1-2) Man a1- Gal b1-3 (Glc a1-4) Man a1- Gal b1-3 (Glc a1-6) Man a1- Gal b1-4 (Glc a1-2) Man a1- Gal b1-4 (Glc a1-3) Man a1- Gal b1-4 (Glc a1-6) Man a1- Gal b1-6 (Glc a1-2) Man a1- Gal b1-6 (Glc a1-3) Man a1- Gal b1-6 (Glc a1-4) Man a1- Gal a1-2 (Glc b1-3) Man a1- Gal a1-2 (Glc b1-4) Man a1- Gal a1-2 (Glc b1-6) Man a1- Gal a1-3 (Glc b1-2) Man a1- Gal a1-3 (Glc b1-4) Man a1- Gal a1-3 (Glc b1-6) Man a1- Gal a1-4 (Glc b1-2) Man a1- Gal a1-4 (Glc b1-3) Man a1- Gal a1-4 (Glc b1-6) Man a1- Gal a1-6 (Glc b1-2) Man a1- Gal a1-6 (Glc b1-3) Man a1- Gal a1-6 (Glc b1-4) Man a1- Gal b1-2 (Glc b1-3) Man a1- Gal b1-2 (Glc b1-4) Man a1- Gal b1-2 (Glc b1-6) Man a1- Gal b1-3 (Glc b1-2) Man a1- Gal b1-3 (Glc b1-4) Man a1- Gal b1-3 (Glc b1-6) Man a1- Gal b1-4 (Glc b1-2) Man a1- Gal b1-4 (Glc b1-3) Man a1- Gal b1-4 (Glc b1-6) Man a1- Gal b1-6 (Glc b1-2) Man a1- Gal b1-6 (Glc b1-3) Man a1- Gal b1-6 (Glc b1-4) Man a1- Gal a1-2 (Glc a1-3) Man b1- Gal a1-2 (Glc a1-4) Man b1- Gal a1-2 (Glc a1-6) Man b1- Gal a1-3 (Glc a1-2) Man b1- Gal a1-3 (Glc a1-4) Man b1- Gal a1-3 (Glc a1-6) Man b1- Gal a1-4 (Glc a1-2) Man b1- Gal a1-4 (Glc a1-3) Man b1- Gal a1-4 (Glc a1-6) Man b1- Gal a1-6 (Glc a1-2) Man b1- Gal a1-6 (Glc a1-3) Man b1- Gal a1-6 (Glc a1-4) Man b1- Gal b1-2 (Glc a1-3) Man b1- Gal b1-2 (Glc a1-4) Man b1- Gal b1-2 (Glc a1-6) Man b1- Gal b1-3 (Glc a1-2) Man b1- Gal b1-3 (Glc a1-4) Man b1- Gal b1-3 (Glc a1-6) Man b1- Gal b1-4 (Glc a1-2) Man b1- Gal b1-4 (Glc a1-3) Man b1- Gal b1-4 (Glc a1-6) Man b1- Gal b1-6 (Glc a1-2) Man b1- Gal b1-6 (Glc a1-3) Man b1- Gal b1-6 (Glc a1-4) Man b1- Gal a1-2 (Glc b1-3) Man b1- Gal a1-2 (Glc b1-4) Man b1- Gal a1-2 (Glc b1-6) Man b1- Gal a1-3 (Glc b1-2) Man b1- Gal a1-3 (Glc b1-4) Man b1- Gal a1-3 (Glc b1-6) Man b1- Gal a1-4 (Glc b1-2) Man b1- Gal a1-4 (Glc b1-3) Man b1- Gal a1-4 (Glc b1-6) Man b1- Gal a1-6 (Glc b1-2) Man b1- Gal a1-6 (Glc b1-3) Man b1- Gal a1-6 (Glc b1-4) Man b1- Gal b1-2 (Glc b1-3) Man b1- Gal b1-2 (Glc b1-4) Man b1- Gal b1-2 (Glc b1-6) Man b1- Gal b1-3 (Glc b1-2) Man b1- Gal b1-3 (Glc b1-4) Man b1- Gal b1-3 (Glc b1-6) Man b1- Gal b1-4 (Glc b1-2) Man b1- Gal b1-4 (Glc b1-3) Man b1- Gal b1-4 (Glc b1-6) Man b1- Gal b1-6 (Glc b1-2) Man b1- Gal b1-6 (Glc b1-3) Man b1- Gal b1-6 (Glc b1-4) Man b1- Gal a1-2 (Man a1-3) Glc a1- Gal a1-2 (Man a1-4) Glc a1- Gal a1-2 (Man a1-6) Glc a1- Gal a1-3 (Man a1-2) Glc a1- Gal a1-3 (Man a1-4) Glc a1- Gal a1-3 (Man a1-6) Glc a1- Gal a1-4 (Man a1-2) Glc a1- Gal a1-4 (Man a1-3) Glc a1- Gal a1-4 (Man a1-6) Glc a1- Gal a1-6 (Man a1-2) Glc a1- Gal a1-6 (Man a1-3) Glc a1- Gal a1-6 (Man a1-4) Glc a1- Gal b1-2 (Man a1-3) Glc a1- Gal b1-2 (Man a1-4) Glc a1- Gal b1-2 (Man a1-6) Glc a1- Gal b1-3 (Man a1-2) Glc a1- Gal b1-3 (Man a1-4) Glc a1- Gal b1-3 (Man a1-6) Glc a1- Gal b1-4 (Man a1-2) Glc a1- Gal b1-4 (Man a1-3) Glc a1- Gal b1-4 (Man a1-6) Glc a1- Gal b1-6 (Man a1-2) Glc a1- Gal b1-6 (Man a1-3) Glc a1- Gal b1-6 (Man a1-4) Glc a1- Gal a1-2 (Man b1-3) Glc a1- Gal a1-2 (Man b1-4) Glc a1- Gal a1-2 (Man b1-6) Glc a1- Gal a1-3 (Man b1-2) Glc a1- Gal a1-3 (Man b1-4) Glc a1- Gal a1-3 (Man b1-6) Glc a1- Gal a1-4 (Man b1-2) Glc a1- Gal a1-4 (Man b1-3) Glc a1- Gal a1-4 (Man b1-6) Glc a1- Gal a1-6 (Man b1-2) Glc a1- Gal a1-6 (Man b1-3) Glc a1- Gal a1-6 (Man b1-4) Glc a1- Gal b1-2 (Man b1-3) Glc a1- Gal b1-2 (Man b1-4) Glc a1- Gal b1-2 (Man b1-6) Glc a1- Gal b1-3 (Man b1-2) Glc a1- Gal b1-3 (Man b1-4) Glc a1- Gal b1-3 (Man b1-6) Glc a1- Gal b1-4 (Man b1-2) Glc a1- Gal b1-4 (Man b1-3) Glc a1- Gal b1-4 (Man b1-6) Glc a1- Gal b1-6 (Man b1-2) Glc a1- Gal b1-6 (Man b1-3) Glc a1- Gal b1-6 (Man b1-4) Glc a1- Gal a1-2 (Man a1-3) Glc b1- Gal a1-2 (Man a1-4) Glc b1- Gal a1-2 (Man a1-6) Glc b1- Gal a1-3 (Man a1-2) Glc b1- Gal a1-3 (Man a1-4) Glc b1- Gal a1-3 (Man a1-6) Glc b1- Gal a1-4 (Man a1-2) Glc b1- Gal a1-4 (Man a1-3) Glc b1- Gal a1-4 (Man a1-6) Glc b1- Gal a1-6 (Man a1-2) Glc b1- Gal a1-6 (Man a1-3) Glc b1- Gal a1-6 (Man a1-4) Glc b1- Gal b1-2 (Man a1-3) Glc b1- Gal b1-2 (Man a1-4) Glc b1- Gal b1-2 (Man a1-6) Glc b1- Gal b1-3 (Man a1-2) Glc b1- Gal b1-3 (Man a1-4) Glc b1- Gal b1-3 (Man a1-6) Glc b1- Gal b1-4 (Man a1-2) Glc b1- Gal b1-4 (Man a1-3) Glc b1- Gal b1-4 (Man a1-6) Glc b1- Gal b1-6 (Man a1-2) Glc b1- Gal b1-6 (Man a1-3) Glc b1- Gal b1-6 (Man a1-4) Glc b1- Gal a1-2 (Man b1-3) Glc b1- Gal a1-2 (Man b1-4) Glc b1- Gal a1-2 (Man b1-6) Glc b1- Gal a1-3 (Man b1-2) Glc b1- Gal a1-3 (Man b1-4) Glc b1- Gal a1-3 (Man b1-6) Glc b1- Gal a1-4 (Man b1-2) Glc b1- Gal a1-4 (Man b1-3) Glc b1- Gal a1-4 (Man b1-6) Glc b1- Gal a1-6 (Man b1-2) Glc b1- Gal a1-6 (Man b1-3) Glc b1- Gal a1-6 (Man b1-4) Glc b1- Gal b1-2 (Man b1-3) Glc b1- Gal b1-2 (Man b1-4) Glc b1- Gal b1-2 (Man b1-6) Glc b1- Gal b1-3 (Man b1-2) Glc b1- Gal b1-3 (Man b1-4) Glc b1- Gal b1-3 (Man b1-6) Glc b1- Gal b1-4 (Man b1-2) Glc b1- Gal b1-4 (Man b1-3) Glc b1- Gal b1-4 (Man b1-6) Glc b1- Gal b1-6 (Man b1-2) Glc b1- Gal b1-6 (Man b1-3) Glc b1- Gal b1-6 (Man b1-4) Glc b1- Glc a1-2 (Man a1-3) Gal a1- Glc a1-2 (Man a1-4) Gal a1- Glc a1-2 (Man a1-6) Gal a1- Glc a1-3 (Man a1-2) Gal a1- Glc a1-3 (Man a1-4) Gal a1- Glc a1-3 (Man a1-6) Gal a1- Glc a1-4 (Man a1-2) Gal a1- Glc a1-4 (Man a1-3) Gal a1- Glc a1-4 (Man a1-6) Gal a1- Glc a1-6 (Man a1-2) Gal a1- Glc a1-6 (Man a1-3) Gal a1- Glc a1-6 (Man a1-4) Gal a1- Glc b1-2 (Man a1-3) Gal a1- Glc b1-2 (Man a1-4) Gal a1- Glc b1-2 (Man a1-6) Gal a1- Glc b1-3 (Man a1-2) Gal a1- Glc b1-3 (Man a1-4) Gal a1- Glc b1-3 (Man a1-6) Gal a1- Glc b1-4 (Man a1-2) Gal a1- Glc b1-4 (Man a1-3) Gal a1- Glc b1-4 (Man a1-6) Gal a1- Glc b1-6 (Man a1-2) Gal a1- Glc b1-6 (Man a1-3) Gal a1- Glc b1-6 (Man a1-4) Gal a1- Glc a1-2 (Man b1-3) Gal a1- Glc a1-2 (Man b1-4) Gal a1- Glc a1-2 (Man b1-6) Gal a1- Glc a1-3 (Man b1-2) Gal a1- Glc a1-3 (Man b1-4) Gal a1- Glc a1-3 (Man b1-6) Gal a1- Glc a1-4 (Man b1-2) Gal a1- Glc a1-4 (Man b1-3) Gal a1- Glc a1-4 (Man b1-6) Gal a1- Glc a1-6 (Man b1-2) Gal a1- Glc a1-6 (Man b1-3) Gal a1- Glc a1-6 (Man b1-4) Gal a1- Glc b1-2 (Man b1-3) Gal a1- Glc b1-2 (Man b1-4) Gal a1- Glc b1-2 (Man b1-6) Gal a1- Glc b1-3 (Man b1-2) Gal a1- Glc b1-3 (Man b1-4) Gal a1- Glc b1-3 (Man b1-6) Gal a1- Glc b1-4 (Man b1-2) Gal a1- Glc b1-4 (Man b1-3) Gal a1- Glc b1-4 (Man b1-6) Gal a1- Glc b1-6 (Man b1-2) Gal a1- Glc b1-6 (Man b1-3) Gal a1- Glc b1-6 (Man b1-4) Gal a1- Glc a1-2 (Man a1-3) Gal b1- Glc a1-2 (Man a1-4) Gal b1- Glc a1-2 (Man a1-6) Gal b1- Glc a1-3 (Man a1-2) Gal b1- Glc a1-3 (Man a1-4) Gal b1- Glc a1-3 (Man a1-6) Gal b1- Glc a1-4 (Man a1-2) Gal b1- Glc a1-4 (Man a1-3) Gal b1- Glc a1-4 (Man a1-6) Gal b1- Glc a1-6 (Man a1-2) Gal b1- Glc a1-6 (Man a1-3) Gal b1- Glc a1-6 (Man a1-4) Gal b1- Glc b1-2 (Man a1-3) Gal b1- Glc b1-2 (Man a1-4) Gal b1- Glc b1-2 (Man a1-6) Gal b1- Glc b1-3 (Man a1-2) Gal b1- Glc b1-3 (Man a1-4) Gal b1- Glc b1-3 (Man a1-6) Gal b1- Glc b1-4 (Man a1-2) Gal b1- Glc b1-4 (Man a1-3) Gal b1- Glc b1-4 (Man a1-6) Gal b1- Glc b1-6 (Man a1-2) Gal b1- Glc b1-6 (Man a1-3) Gal b1- Glc b1-6 (Man a1-4) Gal b1- Glc a1-2 (Man b1-3) Gal b1- Glc a1-2 (Man b1-4) Gal b1- Glc a1-2 (Man b1-6) Gal b1- Glc a1-3 (Man b1-2) Gal b1- Glc a1-3 (Man b1-4) Gal b1- Glc a1-3 (Man b1-6) Gal b1- Glc a1-4 (Man b1-2) Gal b1- Glc a1-4 (Man b1-3) Gal b1- Glc a1-4 (Man b1-6) Gal b1- Glc a1-6 (Man b1-2) Gal b1- Glc a1-6 (Man b1-3) Gal b1- Glc a1-6 (Man b1-4) Gal b1- Glc b1-2 (Man b1-3) Gal b1- Glc b1-2 (Man b1-4) Gal b1- Glc b1-2 (Man b1-6) Gal b1- Glc b1-3 (Man b1-2) Gal b1- Glc b1-3 (Man b1-4) Gal b1- Glc b1-3 (Man b1-6) Gal b1- Glc b1-4 (Man b1-2) Gal b1- Glc b1-4 (Man b1-3) Gal b1- Glc b1-4 (Man b1-6) Gal b1- Glc b1-6 (Man b1-2) Gal b1- Glc b1-6 (Man b1-3) Gal b1- Glc b1-6 (Man b1-4) Gal b1-

Major Glycoconjugates of Eukaryotes GlycoproteinsGlycoproteins GlycolipidsGlycolipids ProteoglycansProteoglycans

Major Glycoconjugates of Eukaryotes GlycoproteinsGlycoproteins GlycolipidsGlycolipids ProteoglycansProteoglycans

Glycoproteins N-LinkedN-Linked O-LinkedO-Linked GPI AnchorGPI Anchor O-GlcNAcO-GlcNAc *A non-inclusive list!

Glycoproteins N-LinkedN-Linked O-LinkedO-Linked GPI AnchorGPI Anchor O-GlcNAcO-GlcNAc *A non-inclusive list!

“N-linked” glycan linkage to protein asparagine residues

Families of N-linked glycan structures

Diversity in complex N-linked glycan structure

Examples of structures found as termini on the branches of complex N-linked glycoprotein glycans…. Terminal Saccharide Diversity --- A key to recognition NeuAc  2-6 Gal  1-4 GlcNAc--- NeuAc  2-3 Gal  1-4 GlcNAc--- Gal  1-3 Gal  1-4 GlcNAc--- 4-SO 3 -GalNAc  1-4 GlcNAc--- (-Gal  1-4 GlcNAc  1-3-) n = polylactosamine repeats (n may be >6) (-NeuAc  2-8-) n = polysialic acid (n may be >50) Fuc residues (e.g. Fuc  1-2Gal; Fuc  1-3 GlcNAc; Fuc  1-4 GlcNAc Fuc residues (e.g. Fuc  1-2 Gal; Fuc  1-3 GlcNAc; Fuc  1-4 GlcNAc

Glycoforms add yet more variation-- Whereas a protein is defined by its unique amino acid sequence, the glycan structure at a single protein glycosylation site varies. Although glycan structures vary at each glycosylation site, the profile is reproducible, protein-specific, site- specific, and cell type-specific.

Three glycoforms of CD59

Glycoproteins N-LinkedN-Linked O-LinkedO-Linked GPI AnchorGPI Anchor O-GlcNAcO-GlcNAc *A non-inclusive list!

Glycoproteins N-LinkedN-Linked O-LinkedO-Linked GPI AnchorGPI Anchor O-GlcNAcO-GlcNAc *A non-inclusive list!

“O-linked” glycan linkage to protein serine residues

Examples of variations of O-linked glycans

Glycoproteins N-LinkedN-Linked O-LinkedO-Linked GPI AnchorGPI Anchor O-GlcNAcO-GlcNAc *A non-inclusive list!

Glycoproteins N-LinkedN-Linked O-LinkedO-Linked GPI AnchorGPI Anchor O-GlcNAcO-GlcNAc *A non-inclusive list!

GPI (glycosylphosphatidylinositol) anchored protein

20 kDa protein 3 kDa N-linked glycan 1 kDa O-linked glycan 1.5 kDa GPI anchor plasma membrane CD59, a complement defense glycoprotein

Glycoproteins N-LinkedN-Linked O-LinkedO-Linked GPI AnchorGPI Anchor O-GlcNAcO-GlcNAc *A non-inclusive list!

Glycoproteins N-LinkedN-Linked O-LinkedO-Linked GPI AnchorGPI Anchor O-GlcNAcO-GlcNAc *A non-inclusive list!

O-GlcNAc Note: No further sugar substitutions have been confirmed

 O-GlcNAc is localized to the cytoplasm and nucleus.  O-GlcNAc is present in all higher eukaryotes studied.  O-GlcNAc is as abundant as phosphorylation.  UDP-GlcNAc is as abundant as ATP in some cell types.  O-GlcNAc containing proteins are phosphoproteins.  O-GlcNAc and phosphorylation are often reciprocal.  O-GlcNAc is dynamic, implying a regulatory role. Key Features of O-linked GlcNAc

Three-state model: unmodified / O-GlcNAc / phosphate O-GlcNAc Glycoprotein Unmodified protein Unmodified protein Phosphoprotein OPO 3 2- Phosphatase OGT Phosphatase Kinase O-GlcNAcase Natasha Zachara

Major Glycoconjugates of Eukaryotes GlycoproteinsGlycoproteins GlycolipidsGlycolipids ProteoglycansProteoglycans

Major Glycoconjugates of Eukaryotes GlycoproteinsGlycoproteins GlycolipidsGlycolipids ProteoglycansProteoglycans

A Glycosphingolipid Galactosylceramide (GalCer)

A Ganglioside “GD1a”

Glycolipids are the major glycoconjugates in the brain

Major Glycoconjugates of Eukaryotes GlycoproteinsGlycoproteins GlycolipidsGlycolipids ProteoglycansProteoglycans

Major Glycoconjugates of Eukaryotes GlycoproteinsGlycoproteins GlycolipidsGlycolipids ProteoglycansProteoglycans

Glycosaminoglycans (GAG’s) and Proteoglycans GLYCOSAMINOGLYCANS (GAG’s)GLYCOSAMINOGLYCANS (GAG’s) –long linear glycans made of repeating disaccharides –Hyaluronic acid is the only “stand-alone” GAG, other GAG’s are constituents of … PROTEOGLYCANSPROTEOGLYCANS –GAG’s on proteins –Defined by their repeating disaccharide units –GAG’s on proteoglycans are sulfated

Hyaluronic acid -- a “simple” space filling molecule (GlcU β4 GlcNAc) n

Post-polymerization variations in glycosaminoglycans

Proteoglycans

Cartilage proteoglycan

fibroblast extracellular intracellular Squamus Epithelium Glycoproteins and glycolipids are found prominently on extracellular leaflet of the plasma membrane and on the intralumenal leaflet of lysosomes. They also are found in the ER and Golgi apparatus, where they are synthesized. GAG’s and proteoglycans are found on the cell surface and as prominent structures of the extracelllular matrix.

Glycobiology Introduction to GlycobiologyIntroduction to Glycobiology Glycan structuresGlycan structures Glycan functionsGlycan functions

Glycobiology Introduction to GlycobiologyIntroduction to Glycobiology Glycan structuresGlycan structures Glycan functionsGlycan functions

Glycobiology Introduction to GlycobiologyIntroduction to Glycobiology Glycan structuresGlycan structures Glycan functionsGlycan functions –Recognition –Biophysical effects –Biochemical effects

Glycobiology Introduction to GlycobiologyIntroduction to Glycobiology Glycan structuresGlycan structures Glycan functionsGlycan functions –Recognition –Biophysical effects –Biochemical effects

Glycobiology Introduction to GlycobiologyIntroduction to Glycobiology Glycan structuresGlycan structures Glycan functionsGlycan functions –Recognition General principlesGeneral principles Influenza virusInfluenza virus InflammationInflammation –Biophysical effects –Biochemical effects

Glycobiology Introduction to GlycobiologyIntroduction to Glycobiology Glycan structuresGlycan structures Glycan functionsGlycan functions –Recognition General principlesGeneral principles Influenza virusInfluenza virus InflammationInflammation –Biophysical effects –Biochemical effects

Sharon and Lis (1993) Scientific American Carbohydrate binding determinant Carbohydrate recognition domain (CRD) Glycans in cell-cell recognition Lectin Glycan

Sialoadhesin CRD May, A.P., et al. (1998) Mol. Cell 1,

Principles of glycan- lectin binding: -hydrogen bonds -hydrophobic stacking -multivalent binding -(salt bridges) -(calcium coordination) May, A.P., et al. (1998) Mol. Cell 1,

Sialic acid glycerol carboxylate acetyl Comb & Roseman, J Am Chem Soc 80, 497 (1958) Sialic acids -- Enhanced roles in glycan recognition

Diverse sialic acid glycans at the cell surface Essentials of Glycobiology Second Edition sialic acid

Diverse sialic acid linkages at the cell surface

NeuAc-α2,3-Gal-β1,4-GlcNAc NeuAc-α2,6-Gal-β1,4-GlcNAc

Glycobiology Introduction to GlycobiologyIntroduction to Glycobiology Glycan structuresGlycan structures Glycan functionsGlycan functions –Recognition General principlesGeneral principles Influenza virusInfluenza virus InflammationInflammation –Biophysical effects –Biochemical effects

Life cycle of the influenza virus - the role of sialic acids

Influenza virus hemagglutinin (side and top views, arrows are sugar binding sites)

Sialoglycan structure drives biological recognition: Influenza pandemics are determined by sialic acid linkage specificity Stevens et al (2006) Nat Rev Microbiol. 4:857-64

Influenza virus neuraminidase (sialidase)

Rational design of an anti-influenza drug based on sialic acid binding to the viral neuraminidase

Influenza virus budding in vitro without (top) and with (bottom) neuraminidase inhibitor

Prophylactic efficacy of a sialic acid-based influenza neuraminidase inhibitor Peters, P.H., et al. (2001) J Am Geriatr Soc 49, 1025

Glycobiology Introduction to GlycobiologyIntroduction to Glycobiology Glycan structuresGlycan structures Glycan functionsGlycan functions –Recognition General principlesGeneral principles Influenza virusInfluenza virus InflammationInflammation –Biophysical effects –Biochemical effects

Inflammation: How we fight bacterial infection

Leukocyte adhesion to E-selectin transfected COS cells

Selectin glycoconjugate Endothelium Neutrophil

Leukocyte cell surface glycan ligand required for endothelial recognition R

Marquardt, et al. (1999) J. Pediatr. 134, 681 Leukocyte adhesion deficiency (LAD) Type II

Human blood group antigens and Selectin ligands

“Within days of starting fucose therapy, neutrophil levels returned to the normal range.... there were no further infections and antibiotic prophylaxis was discontinued.” Marquardt, et al. (1999) Blood 94, 3976

Glycobiology Introduction to GlycobiologyIntroduction to Glycobiology Glycan structuresGlycan structures Glycan functionsGlycan functions –Recognition –Biophysical effects –Biochemical effects

Glycobiology Introduction to GlycobiologyIntroduction to Glycobiology Glycan structuresGlycan structures Glycan functionsGlycan functions –Recognition –Biophysical effects –Biochemical effects

Hyaluronic acid -- a “simple” space filling molecule (GlcU β4 GlcNAc) n

Glycobiology Introduction to GlycobiologyIntroduction to Glycobiology Glycan structuresGlycan structures Glycan functionsGlycan functions –Recognition –Biophysical effects –Biochemical effects

Glycobiology Introduction to GlycobiologyIntroduction to Glycobiology Glycan structuresGlycan structures Glycan functionsGlycan functions –Recognition –Biophysical effects –Biochemical effects

Anticoagulant carbohydrate determinant

Reference texts Reference texts Reference texts Reference texts Oxford University Press, New York Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY. Content freely available at:

Introduction to Glycobiology Introduction to Glycobiology ME: Mar-April 2013 Mar-April 2013 Introduction to Glycobiology Introduction to Glycobiology ME: Mar-April 2013 Mar-April 2013 Course co-directors: Ronald Schnaar Ronald Schnaar Natasha Zachara Natasha Zachara

© 2012 The National Academy of Sciences