Review of glycosylation branch (focus on protein glycosylation)

[rozaru]

Currently, it is difficult to understand how the glycosylation branch terms should be used for annotation as several of its terms represent one single MF.
It is also not clear how they fit with glycoprotein metabolic process and glycolipid metabolic process terms.
The way in which protein glycosylation children are organised as well as their definition is confusing.

Current trees:

glycosylation
|__lipid glycosylation
|__macromolecule glycosylation
|   |__protein glycosylation(+)
|__RNA glycosylation
|__mannosylation
|   |__protein mannosylation
|__fucosylation
|   |__N-glycan fucosylation
|   |__protein O-linked fucosylation

carbohydrate derivative metabolic process (partial tree display)
|__glycoprotein metabolic process(+)
|__liposaccharide metabolic process(+)
|   |__glycolipid metabolic process

Proposal:

• Remove the glycosylation branch and relocate the children to more appropriate branch if necessary
• Move RNA glycosylation to be a child of carbohydrate derivative metabolic process
  Maybe rename it glycoRNA metabolic process?
• Obsolete lipid glycosylation term as the definition represents a MF term
  The genes annotated to it could be instead annotated with glycolipid biosynthetic process (manual checking would be required)
• fucosylation and mannosylation terms represent a MF term
  the gene annotated to them would be better annotated to either glycoprotein biosynthetic process or glycolipid biosynthetic process terms
• Obsolete macromolecule glycosylation term as the definition represents a MF term
• Obsolete the term protein glycosylation and add its children under glycoprotein biosynthetic process (for details see below)

Related tickets:
#29735
#27105
#28977

[rozaru]

The protein glycosylation children would need to be moved under glycoprotein biosynthetic process but before children term definitions need to be improved to avoid ambiguities (most are a description of a MF) and fit better with the literature classification:

• classification hierarchy: by atom then by type of 1st sugar added
• definition includes all the steps for the synthesis of the full glycan (initiation, elongation, processing and sugar modification)

Background

The length and composition of glycans attached to proteins vary between species and often between cells and tissues.

Reference:

• PMID: 33087899 (review on protein glycosylation in human, classification)
• O-glycosylation across kingdom

---

1. Classification
The various type of glycan are classified according to the atom the sugar is attached to. A given atom belongs to a subset of aa residues.

• N-glycosylation

  • Asn
  • Arg (only 1 example known: sweet corn glycoprotein, Arg is found in N-linkage to Glc)

• O-glycosylation

  • Ser
  • Thr
  • Tyr
  • hydroxyproline
  • hydroxylysine

• C-glycosylation

  • Trp

• S-glycosylation

  • Cys

• phosphate-linked (?): protein-Ser-P-Glc

They are also classified according to the 1st sugar that is covalently linked to the aa residue (the term usally cover not only the 1st sugar added but also the full glycan made).
They can be further classified according to the type of chain made in some cases.

• N-GlcNAcylation

  • high mannose N-glycan
  • hybrid N-glycan
  • complex N-glycan
  • keratan sulfate proteoglycan

• O-mannosylation

  • core M0, M1, M2 and M3

• O-GlCNAcylation

• O-fucosylation

• O-GalNAcylation

  • core 1 to 8 (mucin-type O-glycosylation)
  • keratan sulfate proteoglycan

• O-xylosylation

  • HS, heparin, DS, CS proteoglycan

• O-glucosylation

• O-arbinosylation (plants)

• O-galactosylation (example:collagen)

• C-mannosylation

---

2. Metabolism
2.1 biosynthesis of glycoprotein

• Steps:
  • an initiation sep: addition of the 1st sugar to the aa residue
  • an elongation step: sequential addition of subsequent sugars
  • a processing step: sugar may be removed to facilitate the addition of other sugars
  • a sugar modification step: the sugar itself can be modified (acetylation, sulfation etc..)
• involve glycosyltransferases, glucosidases, sulfotransferases etc..
• synthesis predominantly occurs in ER and Golgi, but occurs also in cytosol, nucleus and mitochondria

2.2. catabolism of glycoproteins

• involve endo- and exo- glycosidases
• usually occurs in the lysosome

[rozaru]

This is an example showing why the current terms for protein glycosylation children are not straightforward to use.
Main issue: It is not clear if the definition refers to a single step process that is covered by an MF or to the sequential steps to form the glycan attached to the protein.

For example:
in fly, the synthesis of the O-linked fucosylated glycan involves the sequential action of 3 enzymes: O-fut1, fng and an unknown glucuronidase

This is the definition of the BP GO term that would fit the synthesis according to the literature classification:

GO:0036066 protein O-linked fucosylation
The process of transferring a fucosyl group to a serine or threonine residues in a protein acceptor molecule, to form an O-linked protein-sugar linkage.

However, the definition looks like it describes MF peptide-O-fucosyltransferase activity and could only be used to annotate O-fut1.

Looking at the history of fng annotation shows that curators are confused how to use this GO term:
first, the curator used protein O-linked glycosylation to annotate fng
then the same curator 2 days later, probably having thought about the meaning of the definition, changed it to protein glycosylation
also none of fng homologs have an BP annotation that covers protein glycosylation

[rozaru]

Current tree with proposed changes:

protein glycosylation merge with glycoprotein biosynthetic process
|__protein C-linked glycosylation **OK**
|   |__protein C-linked glycosylation via tryptophan **tag:not for direct annotation**
|       |__protein C-linked glycosylation via 2'-alpha-mannosyl-L-tryptophan **tag:not for direct annotation**

|__protein galactosylation (3 EXP) **MF definition obsolete ?**

|__protein glucuronylation (0 EXP) **MF definition obsolete ?**
|   |__N-terminal protein amino acid glucuronylation (0 EXP) **MF definition obsolete ?**
|       |__N-terminal peptidyl-glycine N-glucuronylation (0 EXP) **MF definition obsolete ?** 

|__protein mannosylation **MF definition obsolete-merge?** 
|   |__chain elongation of O-linked mannose residue **merge with protein O-linked mannosylation**
|   |__protein C-linked glycosylation via 2'-alpha-mannosyl-L-tryptophan **tag:not for direct annotation**
|   |__protein O-linked mannosylation **OK MF definition to update to cover the whole O-mannosylated glycan**

|__protein N-linked glycosylation **OK** 
|   |__protein N-linked glycosylation via asparagine **OK MF definition to update to cover the whole N-glycan**
|       |__(part of) dolichol-linked oligosaccharide biosynthetic process(+) **OK** 

|__protein O-linked glycosylation OK
|   |__O-glycan processing **OK change name to protein O-linked GalNAcylation (mucin type)**
|   |   |__core 1 O-glycan biosynthetic process  (O EXP) **(do we want to keep this level of granularity?)**
|   |   |__core 2 O-glycan biosynthetic process  (2 EXP) **(do we want to keep this level of granularity?)**
|   |   |__core 3 O-glycan biosynthetic process  (O EXP) **(do we want to keep this level of granularity?)**

|   |__protein O-linked fucosylation  **OK MF definition to update to cover the whole O-fucosylated glycan**

|   |__protein O-linked glycosylation via hydroxylysine (0 EXP) **obsolete MF definition** 
|   |__protein O-linked glycosylation via hydroxyproline (9 EXP) **obsolete MF definition (some of the annotation could GO to other O-linked [sugar]ylation)**
|   |__protein O-linked glycosylation via serine (20 EXP) **obsolete MF definition (some of the annotation could GO to other O-linked [sugar]ylation)**
|   |__protein O-linked glycosylation via threonine (24 EXP) **obsolete MF definition (some of the annotation could GO to other O-linked [sugar]ylation)**
|   |   |__protein O-GlcNAcylation via threonine (0 EXP) **obsolete MF definition annotation could go to new term protein O-linked GlcNAcylation**
|   |__protein O-linked glycosylation via tyrosine (0 EXP) **obsolete MF definition** 

|   |__protein O-linked mannosylation **OK MF definition to update to cover the whole O-mannosylated glycan**

|__protein phosphate-linked glycosylation (0 EXP) **MF definition ???**
|   |__protein phosphate-linked glycosylation via serine **tag:not for direct annotation**

|__protein S-linked glycosylation OK
|   |__protein S-linked glycosylation via cysteine (1 EXP) **MF definition tag:not for direct annotation**

New proposed tree:

glycoprotein biosynthetic process
|__protein C-linked glycosylation
|__protein S-linked glycosylation
|__protein N-linked glycosylation
|   |__protein N-linked glycosylation via asparagine 
|       |__(part of) dolichol-linked oligosaccharide biosynthetic process(+)
|       |__keratan sulfate biosynthetic process
|__protein O-linked glycosylation 
|   |__protein O-linked mannosylation
|   |__protein O-linked GalNAcylation (mucin-type)
|       |__keratan sulfate biosynthetic process
|   |__protein O-linked fucosylation
|   |__protein O-linked GlcNAcylation
|   |__protein O-linked glucosylation (**NEW)**
|   |__protein O-linked arbinosylation **(NEW)**
|   |__protein O-linked xylosylation **(NEW)**
|   	|__heparan sulfate biosynthetic process
|	|__heparin sulfate biosynthetic process
|	|__chondroitin sulfate biosynthetic process
|	|__dermatan sulfate biosynthetic process
|       |__glycosaminoglycan-protein linkage region biosynthetic process
|   |__protein O-linked galactosylation **(NEW)**
|__protein phosphate-linked glycosylation (0 EXP) MF definition ???

@pgaudet it’s a lot of information but I thought it was important to see the whole branch to get an idea what potentially needs to be done. If people agree I will break it down into small manageable parts.