Lecture 6 O-GalNac Glycans C-type Lectins

The Glycome is Rich

Examples of O-Glycans Ser/Thr GalNAc Mucins Ser/Thr Man Yeast mannoproteins  -dystroglycan Ser/Thr Fuc Notch Coagulation Factors Fibrinolytic Factors Ser/Thr GlcNAc Nuclear Proteins Cytoplasmic Proteins

More O-glycans Ser Glc Proteoglycans (Glycosaminglycans) Ser Xyl Thr Rho proteins Worm Collagens Dictyostelium proteins ManGlcGalGlcNAc

Even more…. HydroxyLys Gal Collagen C1q complement Plant glycoproteins Dictyostelium HydroxyPro AraGalGlcNAc Tyr Glycogenin Glc

O-Glycosidic Linkage  O-glycosidic linkage is sensitive to alkali (regardless of stereochemistry)  -elimination O OH H H HO H O NAc H H OH NH 2 C H C H 2 C O Ser GalNAc :OH

Fritz, Timothy A. et al. (2004) Proc. Natl. Acad. Sci. USA 101, Ribbon drawing of the ppGaNTase-T1 crystal structure

Fritz, Timothy A. et al. (2004) Proc. Natl. Acad. Sci. USA 101, Modeled binding of a MUC1 glycopeptide to ppGaNTase-T1

Core 3 Ser/Thr 33 Core 4 Ser/Thr 33 66 33 Core 1 Ser/Thr 33 66 Core 2 Core 7 Ser/Thr 66 Core 6? Ser/Thr 66 Core 5 Ser/Thr 33 Core 8 Ser/Thr 33 Other Core O-Glycan Structures

Ser/Thr 33 T-antigen Ser/Thr  3GalT Tn-antigen  6GlcNAcT 33 66 Ser/Thr Core 2 33 33 ST3Gal-I, II, IV 33 66 disialyl T-antigen ST6GalNAc III, IV, I, II sialyl Tn- antigen 66 ST6GalNAc-I 33 66 sialyl 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: Ser/Thr Tn-antigen Ser/Thr 33 T-antigen  3GalT

His-tagged Cosmc HPC4 tagged T- synthase Ni+ purification, Western with mAb to HPC4 T-synthase T-synthase + Mut Cosmc T-synthase + Wt Cosmc Total Not bound to Ni+ Bound to Ni+ Is Cosmc a Chaperone?

Outer Chain Assembly Sequential action of  4GalT and  3GlcNAcT gives rise to polylactosamine chains (Type II repeats) Type I repeats (Gal  3GlcNAc  3) also occur GlcNAc  6Gal 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4 33 44 33 44 33 33 33 Ser/Thr 33 6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