N

N

NOST

UDP-

Glycoproteinfolding

ER

O-Glycans

Golgi

Golgi

Blood groups

Tri- and tetra-antennaryLactosamine extensions

Sialylation (NANA)fucosylation

BisectingGlcNAc

Core fucosylation(α1,6)

Blood groupB

NGNA(non-human)

High mannose

Gal-Galepitope

Lewisx

Lex

α3

β3

β4

R

H

R

B

R

A

R

Lewisy

Ley

α3

α2

β3

R

β4

N

Sialyl Lewisx

SLex

α3

α3

β3

β4

R

Lipid-linkedoligosaccharide

UDP-UDP-

UDP-

UDP-

UDP-

GDP-GDP-

GDP-

UDP-

UDP-

UDP-

UDP-

UDP- (NANA)

CMP-

CMP-

( )

Golgi

Lactosamine

Core 1 Core 2

Core 3

GalNAcO-glycans

Core 4

Galα1-3Galantigen

TerminalNGNA

( )

α3

RR

α3/6

R

Polysialicacid

α8

α8

R

TerminalNANA

α3/6

R

R = Any sugar, extensionscommon to N- and O-glycans

( )

( )

N-GlycansS/T

S/T

S/T

S/T

S/T

Synthesis of N-glycans in the ERis common to all eukaryotes

PP

Extensions

• Other ER/Golgi glycans, such as O-glucose, O-fucose, O-mannose, C-mannose, are found only in specific proteins or tissues.

• Membrane proteins may have glycosylphosphatidil inositol (GPI) anchors.• Glycosaminoglycans (GAGs) are abundant in the extracellular matrix.• Cytosolic and nuclear proteins can be modified with O-GlcNAc.

Insects

UDP-

Paucimannose(short mannose glycans)

Core 1,3 and1,6 fucosylation

(antigenic to humans)

Galactose andsialic acid extensions

in lower arm

High mannose

Insect cells synthesize GalNAc O-glycans linked to serine and threonine, similar to mammalian cells.

Golgi

Plants

UDP-

UDP-

Vacuole

Core 1,3 fucosylationxylose side-chain

(antigenic to humans)

High mannose

Paucimannose

Complex

Unique plant O-glycans are arabinose or galactose residues linked to hydroxyproline, abundant in structural glycoproteins of the cell wall. Plants also have GalNAc O-glycans linked to serine and threonine.

Golgi

Yeasts

GDP-

GDP-

GDP-

Mannan (cell wall proteins)hypermannosylation

β-mannosechains

S. cerevisiase

Pichia

K. lactis

UDP-

UDP-

K. lactis

[ ]P

+

+

+

[ ]P

n

][n

][n

Linear short chains of mannose residues, sometimes terminated with galactose, are the most common O-glycan modifications in yeasts.

Golgi

TerminalGlcNAc residues

Deglycosylation Enzymes• NEW – Rapid PNGase F• PNGase F & PNGase F (Glycerol-free)• PNGase F & PNGase F (GF), Recombinant• Protein Deglycosylation Mix• Endo H & Endo Hf

• O-Glycosidase• Remove-iT® Endo S• Remove-iT Endo D• Remove-iT PNGase F

Exoglycosidase Enzymes• α-N-Acetylgalactosaminidase• β-N-Acetylglucosaminidase• β-N-Acetylhexosaminidasef

Exoglycosidase Enzymes (cont’d)

• β-N-Acetylglucosaminidase S• α1-2 Fucosidase• α1-2,4,6 Fucosidase• α1-3,6 Galactosidase• α1-3,4,6 Galactosidase• β1-3 Galactosidase• β1-4 Galactosidase• β1-4 Galactosidase S• α1-2,3 Mannosidase• α1-6 Mannosidase• α2-3,6,8 Neuraminidase• α2-3,6,8,9 Neuraminidase A• α2-3 Neuraminidase• α2-3 Neuraminidase S

Glycoproteomics• Trypsin-digested BSA MS Standard

(CAM Modified)• Trypsin-ultra™, Mass Spectrometry Grade• Endoproteinase GluC• Endoproteinase AspN• Proteinase K, Molecular Biology Grade

Heparin Lyase Enzymes• Bacteroides Heparinase I• Bacteroides Heparinase II• Bacteroides Heparinase III

Other Related Products• K. lactis Expression Kit• RNase B Standard• Fetuin Glycoprotein Standard

A wide variety of glycan species are found in secretory and membrane glycoproteins

Secretion or localization to plasma membrane

Secretion of glycoproteins with high-mannose glycans

Mannose-phosphatevacuole localization

PROTEIN MODIFICATIONS IN DIFFERENT EXPRESSION SYSTEMSGLYCOSYLATION

PRODUCTS FROM NEW ENGLAND BIOLABS®

Mammalian cells

Gal GalNacMan GlcNAc NeuAc (NANA) NeuGc (NGNA)Fuc XylGlc

NEW ENGLAND BIOLABS®, NEB® and REMOVE-IT® are registered trademarks of New England Biolabs, Inc. TRYPSIN-ULTRA™ is a trademark of New England Biolabs, Inc. | v2.0 5/15

To view online tutorials, learn more about products available from NEB, or to download our Glycobiology Brochure, visit NEBGlycosidase.com.

P = Phosphate residueRef: Varki, et al. Essentials of Glycobiology, Second Edition. New York: Cold Spring Harbor Laboratory Press, 2009.