The gut microbiota harbor diverse β-glucuronidase (GUS) enzymes that liberate glucuronic acid (GlcA) sugars from small-molecule conjugates and complex carbohydrates. However， only the Enterobacteriaceae family of human gut-associated Proteobacteria maintain a GUS operon under the transcriptional control of a glucuronide repressor， GusR. Despite its potential importance in ， ， ， ， and opportunistic pathogens， the structure of GusR has not been examined. Here， we explore the molecular basis for GusR-mediated regulation of GUS expression in response to small-molecule glucuronides. Presented are 2.1-?-resolution crystal structures of GusRs from and in complexes with a glucuronide ligand. The Gus... More
The gut microbiota harbor diverse β-glucuronidase (GUS) enzymes that liberate glucuronic acid (GlcA) sugars from small-molecule conjugates and complex carbohydrates. However， only the Enterobacteriaceae family of human gut-associated Proteobacteria maintain a GUS operon under the transcriptional control of a glucuronide repressor， GusR. Despite its potential importance in ， ， ， ， and opportunistic pathogens， the structure of GusR has not been examined. Here， we explore the molecular basis for GusR-mediated regulation of GUS expression in response to small-molecule glucuronides. Presented are 2.1-?-resolution crystal structures of GusRs from and in complexes with a glucuronide ligand. The GusR-specific DNA operator site in the regulatory region of the GUS operon is identified， and structure-guided GusR mutants pinpoint the residues essential for DNA binding and glucuronide recognition. Interestingly， the endobiotic estradiol-17-glucuronide and the xenobiotic indomethacin-acyl-glucuronide are found to exhibit markedly differential binding to these GusR orthologs. Using structure-guided mutations， we are able to transfer GusR's preferential DNA and glucuronide binding affinity to GusR. Structures of putative GusR orthologs from GUS-encoding Firmicutes species also reveal functionally unique features of the Enterobacteriaceae GusRs. Finally， dominant-negative GusR variants are validated in cell-based studies. These data provide a molecular framework toward understanding the control of glucuronide utilization by opportunistic pathogens in the human gut.
