About the Repository
Human glycosylation enzymes are difficult to produce recombinantly. The Repository was built to remove that obstacle by providing ready-made expression constructs, with the design rationale and sequence of each one on record.
Why these enzymes are hard to express
Mammalian enzymes of glycan synthesis and catabolism — glycosyltransferases (GTs), glycoside hydrolases (GHs) and a range of glycan-modifying enzymes — are synthesised in the endoplasmic reticulum and act on glycans in the lumenal compartments of the secretory and endocytic pathways. They are typically glycosylated, disulfide-bonded and otherwise post-translationally modified, which makes them poor candidates for functional expression in bacteria.
The Repository addresses this by supplying constructs for expression in bacteria, insect cells (baculovirus) and mammalian cells, designed to produce soluble catalytic domains wherever the enzyme's topology allows. Most constructs are truncated to remove the transmembrane segment and fused to affinity tags or larger fusion partners that aid detection, quantitation and purification. All coding regions were captured as Gateway® entry clones in pDONR221 or equivalent synthesised donors, then transferred into custom destination vectors. See Construct Design for the full strategy.
Scope of the collection
- 200 glycosyltransferase records across the CAZy GT families.
- 77 glycoside hydrolase records across the CAZy GH families.
- 62 records for other glycan-related enzymes — sulfotransferases, sulfatases, phosphotransferases, ER lectins and chaperones, protein disulfide isomerases, and enzymes of dolichol-phosphate sugar synthesis — grouped by function because CAZy does not classify them.
- 47 bacterial homologues of mammalian GTs and GHs, chosen as more tractable surrogates and expressed in E. coli.
Research groups
The project was developed through an ARRA supplement to an NCRR research grant to the University of Georgia entitled "Development of a Repository for Glycan-related Enzymes". Four groups contributed:
| Role | Contact | Institution |
|---|---|---|
| Construct design and mammalian cell expression | Kelley Moremen | University of Georgia |
| Construct generation and distribution | Josh LaBaer (Jason Steel, Production Manager) | Arizona State University |
| Baculovirus / insect cell expression | Don Jarvis | University of Wyoming |
| Bacterial expression | Harry Gilbert | University of Georgia (now at Newcastle University) |
Citation
The methods paper for the collection is Moremen et al. (2018) Expression system for structural and functional studies of human glycosylation enzymes, Nature Chemical Biology 14, 156–162 (PMID 29251719). A full list is on the Publications page.
About this version of the site
This site is a rebuild of the original Dreamweaver frameset at https://glycoenzymes.ccrc.uga.edu/. The content is the same set of records; the structure has been rebuilt as standalone, linkable pages generated from a single structured dataset. Practical consequences:
- Every page has its own URL and can be bookmarked, cited or sent to a colleague. In the frameset version the address bar showed only the site root regardless of what was on screen.
- The three separate alphabetical lists are replaced by one sortable, filterable table that also covers the bacterial homologues, which previously had no alphabetical index.
- A search page covers all records at once; the original site had no search.
- Construct availability is stated in words on every row rather than signalled only by a green highlight, so it survives printing and is readable by screen readers.
- The layout is responsive and the pages print sensibly.
- Records are unchanged from the legacy site and carry its annotations as recorded at the time. They have not been re-verified against current database releases.