Structure-function and engineering of plant UDP-glycosyltransferase

Functional Characterization and Protein Engineering of a Glycosyltransferase GcCGT to Produce Flavone 6,8-Di-C- and 6-C-4′-O-Glycosides

“…The GcCGT-apo crystal contains four GcCGT molecules in an asymmetric structural unit, and their structures were basically the same. The structure of GcCGT belongs to the typical GT-B type, consisting of two flexible Rossmann-like β/α/β domains . Subsequently, UDP-Glc was docked into GcCGT based on previously reported structures (Figure S80), including GgCGT/UDP-Glc (PDB ID: 6L5P), SbCGTb/UDP-Glc (PDB ID: 6LFZ), and MiCGT/UDP-Glc (PDB ID: 7VA8).…”

“…The structure of GcCGT belongs to the typical GT-B type, consisting of two flexible Rossmann-like β/ α/β domains. 27 Subsequently, UDP-Glc was docked into GcCGT based on previously reported structures (Figure S80), including GgCGT/UDP-Glc (PDB ID: 6L5P), SbCGTb/UDP-Glc (PDB ID: 6LFZ), and MiCGT/UDP-Glc (PDB ID: 7VA8). 3,6,8 Through structural analysis, we found that F387 was located in a loop (loop 1) near the substrate.…”

“…These two peptides were located at the entrance of the substrate binding pocket (Figure 6D) and exhibited outward-expansion conformations at different simulation times (Figure S81). Hence, the F387K mutant probably has a larger pocket to accommodate orientin (27) as the substrate. Molecular docking showed that orientin (27) could be well docked in the structure of F387K, and the 8-C-glucosyl group of orientin (27) was exactly located near K387 (Figure 6E).…”

“…Therefore, we proposed to modify GcCGT to catalyze flavone 8-C-glycosides (Figure 3A,B). Homology modeling and molecular docking were conducted, and orientin (27) failed to dock into GcCGT/UDP-Glc (Supporting Information). Instead, its aglycone luteolin (1) was docked into GcCGT.…”

Directed Evolution of the UDP-Glycosyltransferase UGTBL1 for Highly Regioselective and Efficient Biosynthesis of Natural Phenolic Glycosides

“…However, the diversity of glycosylation sites leads to variations in the biological activities and functions of glycoside products . When polyphenols are used as aglycone acceptors, chemical glycosylation requires cumbersome protection/deprotection steps to achieve high regioselectivity. , Therefore, the enzymatic glycosylation of polyphenols offers numerous advantages, including precise tuning of regioselectivity, adherence to green chemistry principles, and high efficiency. − UGTs from various species, including Arabidopsis thaliana, Vitis vinifera, and Streptomyces antibioticus, have been extensively characterized. − Indeed, most of the characterized UGTs, including UGT BL 1, exhibit poor regioselectivity and catalytic activity. Simultaneous improvements in the catalytic efficiency and regioselectivity of UGTs remain a challenge.…”