Structural characterization of a pseudaminic acid-modified lateral flagellar filament from Vibrio alginolyticus

Bacterial flagellar filaments are typically modified with glycans, but their structural basis and functional importance in certain species, such as Vibrio alginolyticus, remain elusive. In V. alginolyticus, lateral flagella are responsible for surface-associated motility in viscous environments, with the protein LafA responsible for forming the filament.

A gene encoding a putative flagellin glycosylation factor, maf, is found immediately downstream of lafA, indicating that the lateral filament may be glycosylated. To explore this hypothesis, researchers purified the lateral flagellar filament from V. alginolyticus and utilized electron cryomicroscopy to determine its structure at a resolution of 2.37 [A].

By incorporating the LaF A structure into the map, additional densities were observed near five serine residues, potentially indicating O-linked pseudaminic acid modifications. Mass spectrometry confirmed these modifications as pseudaminic acid attached to Ser148, Ser173, Ser183, Ser189, and Ser197. Further investigation revealed that deletion of the maf gene prevented lateral flagella formation and motility, but complementation with maf restored these functions.

These findings provide the first structural insight into the pseudaminic acid modification of the Vibrio lateral flagellar filament and establish a foundation for understanding the role of flagellin glycosylation in surface-associated motility.