New Paradigms of Pilus Assembly Mechanisms in Gram-Positive Actinobacteria

Adhesive pili in Gram-positive bacteria represent a variety of extracellular multiprotein polymers that mediate bacterial colonization of specific host tissues and associated pathogenesis. Pili are assembled in two distinct but coupled steps, an orderly crosslinking of pilin monomers and subsequent anchoring of the polymer to peptidoglycan, catalyzed by two transpeptidase enzymes – the pilus-specific sortase and the housekeeping sortase. Here, we review this biphasic assembly mechanism based on studies of two prototypical models, the heterotrimeric pili in Corynebacterium diphtheriae and the heterodimeric pili in Actinomyces oris, highlighting some newly emerged basic paradigms. The disparate mechanisms of protein ligation mediated by the pilus-specific sortase and the spatial positioning of adhesive pili on the cell surface modulated by the housekeeping sortase are among the notable highlights. Covalently linked pili are assembled on the cell surface of many Gram-positive bacteria via a biphasic mechanism whereby pilus polymerization is catalyzed by the pilus-specific sortase followed by cell wall anchoring of pili by the housekeeping sortase.Pilus-mediated adhesion depends on pilus length, which is modulated by the housekeeping sortase via unique structural features.Some Gram-positive surface proteins with the LPXTG motif may hijack a pilus assembly machine via molecular mimicry to be displayed at the pilus tip.Pilus-specific sortase enzymes provide a bioconjugation tool via the formation of an isopeptide bond that is mechanically stable and less susceptible to proteolytic cleavage.