Lecture 6

O-GalNac GlycansC-type Lectins

The Glycome is Rich

Examples of O-Glycans

Ser/Thr

GalNAc

Mucins

Ser/Thr

Man

Yeast mannoproteins-dystroglycan

Ser/Thr

Fuc

NotchCoagulation FactorsFibrinolytic Factors

Ser/Thr

GlcNAc

Nuclear ProteinsCytoplasmic Proteins

More O-glycans

Ser

Glc

Proteoglycans(Glycosaminglycans)

Ser

Xyl

Thr

Rho proteinsWorm CollagensDictyostelium proteins

Man Glc Gal GlcNAc

Even more….

HydroxyLys

Gal

CollagenC1q complement

Plant glycoproteinsDictyostelium

HydroxyPro

Ara Gal GlcNAc

Tyr

Glycogenin

Glc

O-Glycosidic Linkage

O-glycosidic linkage is sensitive to alkali (regardless of stereochemistry)

-elimination

O

OH

H

H

HO

H

O

NAcHH

OH

NH2CH

C

H2C

OSer

GalNAc

:OH

Fritz, Timothy A. et al. (2004) Proc. Natl. Acad. Sci. USA 101, 15307-15312

Ribbon drawing of the ppGaNTase-T1 crystal structure

Fritz, Timothy A. et al. (2004) Proc. Natl. Acad. Sci. USA 101, 15307-15312

Modeled binding of a MUC1 glycopeptide to ppGaNTase-T1

Core 3

Ser/Thr

3

Core 4

Ser/Thr

3 6

Ser/Thr

3

Core 1

Ser/Thr

3 6

Core 2

Core 7

Ser/Thr

6

Core 6?

Ser/Thr

6

Core 5

Ser/Thr

3

Core 8

Ser/Thr

3

Other Core O-Glycan Structures

Ser/Thr

3

T-antigen

Ser/Thr

3GalT

Tn-antigen

6GlcNAcT 3 6

Ser/Thr

Core 2

3

3

ST3Gal-I, II, IV

36

disialylT-antigen

ST6GalNAcIII, IV, I, II

sialyl Tn-

antigen6

ST6GalNAc-I

3 6sialyl

T-antigen

ST6GalNAc-II, I

Cosmc

• Tn-antigens accumulate due to loss of 3Gal transferase activity

• No mutations in 3GalT; message is expressed

• Missing Cosmc (Core 1 3GalT -specific molecular chaperone, Xq23

• 25% identity, >40% homology to 3GalT

• Absence of Cosmc results in proteosome degradation of 3GalT

Cosmc

Ju & Cummings (2002) PNAS 99:16613-8

Ser/Thr

Tn-antigen

Ser/Thr

3

T-antigen

3GalT

His-tagged Cosmc

HPC4 tagged T-synthase

Ni+ purification, Western with mAb to HPC4

T-syn

thas

eT-s

ynth

ase

+

Mut C

osmc

T-syn

thas

e +

Wt C

osmc

To

tal

No

t b

ou

nd

to

Ni+

Bo

un

d t

o N

i+

Is Cosmc a Chaperone?

Outer Chain Assembly

• Sequential action of 4GalT and 3GlcNAcT gives rise to polylactosamine chains (Type II repeats)

• Type I repeats (Gal3GlcNAc3) also occur

• GlcNAc6Gal branches (I-antigen) can occur

• The ends of the chains are capped in -linked sugars, e.g. 3/4Fuc and 3/6sialic acids

• Terminal structures make up important blood group determinants, e.g. the Lewis antigens

4

3

4

3

4

3

3

3

Ser/Thr

3 6

Intestinal Epithelium stained with Periodic acid-Schiff’s Reagent

Mucins

• 11p15 family (MUC2, MUC5AC, MUC5B, MUC6) probably responsible for the formation of mucus layers

• 7q22 family (MUC3A, MUC3B, MUC12), 1q21 (MUC1), and 3q (MUC4, MUC13) are membrane mucins

• Mucin families are not necessairly related to each other, convergent evolution

Dekker et al. (2002) TIBS 27:126

C-type Lectins

Leukocyte Rolling

Tim Springer, Harvard