Featured Article - December 2011
Short description: The structures of four glycosyltransferases operating within a single biosynthetic pathway point to conserved and divergent mechanisms for controlling the regioselective addition of sugar donors, with implications for glycosyltransferase engineering.
Engineering glycosyltransferases to accommodate sugar analogs offers opportunities to broaden or alter the utility of an antibacterial or anticancer agent. However, the diversity of both natural product scaffolds and glycan donors used by glycosyltransferases has stymied the development of general rules for enzyme function and redesign strategies. Towards that end, Phillips, Thorson and colleagues (PSI CESG) now report four glycosyltransferase structures that highlight commonalities and differences determining sugar recognition and regioselective activity.
In calicheamicin biosynthesis, four glycosyltransferases are responsible for synthesizing a branched trisaccharide extending from the enediyne scaffold (performed by CalG3, CalG2 and CalG4) and decorating an aromatic group appended to the disaccharide (CalG1). Thus, each enzyme works on the same core molecule, but must attach a carbohydrate to different positions.
The new structures include CalG1, CalG2, and CalG3 with substrates and/or products as well as apo CalG4, which was used to predict the bound structure. Analysis of these structures points to conserved features in binding to the enediyne scaffold (with CalG1 providing some exceptions), including aromatic residues that contact the enediyne itself, a hydrophobic pocket postulated to protect an unusual methylated trisulfide group, and hydrogen bonding to a partner with unused hydroxyl groups. Additionally, a His residue is poised to play the major catalytic role, except in CalG2, which may use a Thr to catalyze the carbohydrate attachment to the more reactive hydroxylamine substrate.
A comparison of the structures identifies three ways in which the proteins diverge to control the reaction product. The first mechanism, using a short or long helix adjacent to the active site, yields different architectures within the protein as a whole to create different active sites. The second mechanism, reflecting a continuous or bent helix near the active site, changes the size of the active site. Finally, the different placement of the catalytic residue, as in CalG2, offers an alternative way of reshaping the reaction without substantial remodeling.
Overall, the combined analysis across four biosynthetically linked structures provides new insights that will shape our understanding of these interesting enzymes and the molecules they produce.
A. Chang et al. Complete set of glycosyltransferase structures in the calicheamicin biosynthetic pathway reveals the origin of regiospecificity.
Proc. Natl. Acad. Sci. 108, 17649-17654 (2011). doi:10.1073/pnas.1108484108