9YQ5

Chlorella virus hyaluronan synthase bound to an inserted UDP-GlcA

Summary for 9YQ5

• Entry DOI: 10.2210/pdb9yq5/pdb
• EMDB information: 73324
• Descriptor: Hyaluronan synthase, nanobody Nb872, nanobody Nb881, ... (6 entities in total)
• Functional Keywords: glycosyltransferase, membrane protein, hyaluronan synthase, substrate, transferase-immune system complex, transferase/immune system
• Biological source: Paramecium bursaria Chlorella virus CZ-2; More
• Total number of polymer chains: 3
• Total formula weight: 96805.58

Authors

Stephens, Z.,Zimmer, J. (deposition date: 2025-10-15, release date: 2025-12-10)

Primary citation

Stephens, Z.,Karasinska, J.,Zimmer, J.
Insights into substrate binding and utilization by hyaluronan synthase.
Biorxiv, 2025

PubMed Abstract: Hyaluronan (HA) is an essential polysaccharide of the vertebrate extracellular matrix. It serves as an adhesive, lubricant, signaling molecule, and spatial filler without which embryogenesis would not complete. HA is synthesized by a membrane-integrated glycosyltransferase (HAS) that polymerizes UDP-activated N-acetylglucosamine and glucuronic acid (GlcA) in an alternating fashion. The nascent HA chain is secreted across the plasma membrane during this process. How HAS couples these tasks remains poorly understood. Here, we employ a combination of structural biology, biochemistry and glycobiology to delineate how HAS recognizes and utilizes UDP-GlcA. Using single-particle cryo-EM, we reveal a two-step process by which HAS binds its substrate UDP-GlcA. Prior to proper insertion into the catalytic pocket, the substrate is bound in a proofreading pose that may increase substrate selectivity. This state is accompanied by conformational changes of active site residues surrounding the UDP-binding pocket and involves a pair of basic residues that sense the substrate's carboxyl group. Further, we establish that HAS is unable to catalyze UDP-GlcA turnover in the absence of an acceptor sugar, emphasizing the role of a priming GlcNAc in glycosyl transfer. Lastly, cryo-EM snapshots of a dodecylmaltoside molecule trapped in the active site provide novel insights into substrate promiscuity. Here, our studies demonstrate that HAS catalyzes semi-selective GlcA-transfer to non-canonical β-linked acceptors.

PubMed: 41280022
DOI: 10.1101/2025.10.17.683186

Experimental method

ELECTRON MICROSCOPY (2.8 Å)