A multi-enzyme machine polymerizes the Haemophilus influenzae type b capsule

• Published in: Nature chemical biology Vol. 19; no. 7; pp. 865 - 877
• Main Authors: Cifuente, Javier O., Schulze, Julia, Bethe, Andrea, Di Domenico, Valerio, Litschko, Christa, Budde, Insa, Eidenberger, Lukas, Thiesler, Hauke, Ramón Roth, Isabel, Berger, Monika, Claus, Heike, D’Angelo, Cecilia, Marina, Alberto, Gerardy-Schahn, Rita, Schubert, Mario, Guerin, Marcelo E., Fiebig, Timm
• Format: Journal Article
• Language: English
• Published: New York Nature Publishing Group US 01-07-2023; Nature Publishing Group
• Subjects: 631/326/590; 631/45/173; 631/45/221; 631/45/72; 631/535/1266; Antigens; Bacteria; Bacterial Capsules - metabolism; Biochemical Engineering; Biochemistry; Bioorganic Chemistry; Biosynthesis; Catalysis; Cell Biology; Chemical synthesis; Chemistry; Chemistry and Materials Science; Chemistry/Food Science; Child; Crystal structure; Enzymes; Fermentation; Glycan; Gram-Negative Bacteria; Haemophilus Infections - microbiology; Haemophilus Infections - prevention & control; Haemophilus influenzae; Haemophilus influenzae type b; Haemophilus Vaccines - metabolism; Host-pathogen interactions; Humans; Infant; NMR; Nuclear magnetic resonance; Pathogens; Polymers; Vaccine development; Vaccines
• ISSN: 1552-4450; 1552-4469
• DOI: 10.1038/s41589-023-01324-3

Bacterial capsules have critical roles in host-pathogen interactions. They provide a protective envelope against host recognition, leading to immune evasion and bacterial survival. Here we define the capsule biosynthesis pathway of
Haemophilus influenzae
serotype b (Hib), a Gram-negative bacterium that causes severe infections in infants and children. Reconstitution of this pathway enabled the fermentation-free production of Hib vaccine antigens starting from widely available precursors and detailed characterization of the enzymatic machinery. The X-ray crystal structure of the capsule polymerase Bcs3 reveals a multi-enzyme machine adopting a basket-like shape that creates a protected environment for the synthesis of the complex Hib polymer. This architecture is commonly exploited for surface glycan synthesis by both Gram-negative and Gram-positive pathogens. Supported by biochemical studies and comprehensive 2D nuclear magnetic resonance, our data explain how the ribofuranosyltransferase CriT, the phosphatase CrpP, the ribitol-phosphate transferase CroT and a polymer-binding domain function as a unique multi-enzyme assembly.
Polymerase Bcs3, which allows the fermentation-free synthesis of
Haemophilus influenzae
type b capsule for vaccine development, adopts a basket-like shape with all six active sites facing the interior, creating a protected environment for catalysis.