Format

Send to

Choose Destination
Mol Microbiol. 2019 Jan 1. doi: 10.1111/mmi.14197. [Epub ahead of print]

Streptococcal dTDP-L-rhamnose biosynthesis enzymes: functional characterization and lead compound identification.

Author information

1
Department of Medical Microbiology, University Medical Center Utrecht, Utrecht University, Heidelberglaan 100, 3584 CX, Utrecht, The Netherlands.
2
Division of Molecular Microbiology, School of Life Sciences, University of Dundee, Dow Street, DD1 5EH, Dundee, UK.
3
Department of Chemistry, Complex Carbohydrate Research Center, The University of Georgia, 315 Riverbend Road, Athens, USA.
4
Mycobacterial Metabolism and Antibiotic Research Laboratory, The Francis Crick Institute, London, UK.
5
Department of Medical Chemistry and Chemical Biology, Utrecht Institute Pharmaceutical Science, University Utrecht, Utrecht, 3508 TB, The Netherlands.

Abstract

Biosynthesis of the nucleotide sugar precursor dTDP-L-rhamnose is critical for the viability and virulence of many human pathogenic bacteria, including Streptococcus pyogenes (Group A Streptococcus; GAS), Streptococcus mutans and Mycobacterium tuberculosis. Streptococcal pathogens require dTDP-L-rhamnose for the production of structurally similar rhamnose polysaccharides in their cell wall. Via heterologous expression in S. mutans, we confirmed that GAS RmlB and RmlC are critical for dTDP-L-rhamnose biosynthesis through their action as dTDP-glucose-4,6-dehydratase and dTDP-4-keto-6-deoxyglucose-3,5-epimerase enzymes respectively. Complementation with GAS RmlB and RmlC containing specific point mutations corroborated the conservation of previous identified catalytic residues. Bio-layer interferometry was used to identify and confirm inhibitory lead compounds that bind to GAS dTDP-rhamnose biosynthesis enzymes RmlB, RmlC and GacA. One of the identified compounds, Ri03, inhibited growth of GAS, other rhamnose-dependent streptococcal pathogens as well as M. tuberculosis with an IC50 of 120-410 µM. Importantly, we confirmed that Ri03 inhibited dTDP-L-rhamnose formation in a concentration-dependent manner through a biochemical assay with recombinant rhamnose biosynthesis enzymes. We therefore conclude that inhibitors of dTDP-L-rhamnose biosynthesis, such as Ri03, affect streptococcal and mycobacterial viability and can serve as lead compounds for the development of a new class of antibiotics that targets dTDP-rhamnose biosynthesis in pathogenic bacteria.

PMID:
30600561
DOI:
10.1111/mmi.14197

Supplemental Content

Full text links

Icon for Wiley
Loading ...
Support Center