5C71
The structure of Aspergillus oryzae a-glucuronidase complexed with glycyrrhetinic acid monoglucuronide
Summary for 5C71
| Entry DOI | 10.2210/pdb5c71/pdb |
| Related | 5c70 |
| Descriptor | Glucuronidase, (3BETA,5BETA,14BETA)-3-HYDROXY-11-OXOOLEAN-12-EN-29-OIC ACID, alpha-D-glucopyranuronic acid, ... (4 entities in total) |
| Functional Keywords | beta-glucuronidase; glycyrrhetinic acid monoglucuronide, hydrolase |
| Biological source | Aspergillus oryzae |
| Total number of polymer chains | 4 |
| Total formula weight | 286311.08 |
| Authors | |
| Primary citation | Lv, B.,Sun, H.,Huang, S.,Feng, X.,Jiang, T.,Li, C. Structure-guided engineering of the substrate specificity of a fungal beta-glucuronidase toward triterpenoid saponins. J.Biol.Chem., 293:433-443, 2018 Cited by PubMed Abstract: Glycoside hydrolases (GHs) have attracted special attention in research aimed at modifying natural products by partial removal of sugar moieties to manipulate their solubility and efficacy. However, these modifications are challenging to control because the low substrate specificity of most GHs often generates undesired by-products. We previously identified a GH2-type fungal β-glucuronidase from (GUS) exhibiting promiscuous substrate specificity in hydrolysis of triterpenoid saponins. Here, we present the GUS structure, representing the first structure of a fungal β-glucuronidase, and that of an inactive GUS mutant in complex with the native substrate glycyrrhetic acid 3--mono-β-glucuronide (GAMG). GUS displayed a homotetramer structure with each monomer comprising three distinct domains: a sugar-binding, an immunoglobulin-like β-sandwich, and a TIM barrel domain. Two catalytic residues, Glu and Glu, acted as acid/base and nucleophile, respectively. Structural and mutational analyses indicated that the GAMG glycan moiety is recognized by polar interactions with nine residues (Asp, His, Asp, Tyr, Tyr, Asp, Arg, Asn, and Lys) and that the aglycone moiety is recognized by aromatic stacking and by a π interaction with the four aromatic residues Tyr, Phe, Trp, and Tyr Finally, structure-guided mutagenesis to precisely manipulate GUS substrate specificity in the biotransformation of glycyrrhizin into GAMG revealed that two amino acids, Ala and Arg, are critical for substrate specificity. Moreover, we obtained several mutants with dramatically improved GAMG yield (>95%). Structural analysis suggested that modulating the interaction of β-glucuronidase simultaneously toward glycan and aglycone moieties is critical for tuning its substrate specificity toward triterpenoid saponins. PubMed: 29146597DOI: 10.1074/jbc.M117.801910 PDB entries with the same primary citation |
| Experimental method | X-RAY DIFFRACTION (2.62 Å) |
Structure validation
Download full validation report
