The basal and major pilins in the Corynebacterium diphtheriaeSpaA pilus adopt similar structures that competitively react with the pilin polymerase

Abstract

Many species of pathogenic gram‐positive bacteria display covalently crosslinked protein polymers (called pili or fimbriae) that mediate microbial adhesion to host tissues. These structures are assembled by pilus‐specific sortase enzymes that join the pilin components together via lysine‐isopeptide bonds. The archetypal SpaA pilus from Corynebacterium diphtheriae is built by the CdSrtA pilus‐specific sortase, which crosslinks lysine residues within the SpaA and SpaB pilins to build the shaft and base of the pilus, respectively. Here, we show that CdSrtA crosslinks SpaB to SpaA via a K139(SpaB)‐T494(SpaA) lysine‐isopeptide bond. Despite sharing only limited sequence homology, an NMR structure of SpaB reveals striking similarities with the N‐terminal domain of SpaA (NSpaA) that is also crosslinked by CdSrtA. In particular, both pilins contain similarly positioned reactive lysine residues and adjacent disordered AB loops that are predicted to be involved in the recently proposed “latch” mechanism of isopeptide bond formation. Competition experiments using an inactive SpaB variant and additional NMR studies suggest that SpaB terminates SpaA polymerization by outcompeting NSpaA for access to a shared thioester enzyme–substrate reaction intermediate.

The crystal structure of the N-terminal domain of the backbone pilin LrpA reveals a new closure-and-twist motion for assembling dynamic pili in Ligilactobacillus ruminis

“…6c and 6d). This structural phenomenon was also observed in GG-SpaD and other backbone pilins (Chaurasia et al, 2018), as well as in the basal pilins from L. rhamnosus GG and Corynebacterium diphtheriae (Megta et al, 2019(Megta et al, , 2020Sue et al, 2023). We compared the N-domains of LrpA FT and LrpA FO with those of the backbone and basal pilin crystal structures from L. rhamnosus GG (Fig.…”

“…While what dictates the length of the pilus is still under debate, the polymerization process appears to terminate when the basal pilin is incorporated at the pilus base Chang et al, 2019). The N-terminal domain of the basal pilin resembles that of backbone pilins, containing a pilin motif, which enables the incorporation of the basal subunit at the bottom of the polymeric pilus structure (Krishnan et al, 2007;Megta et al, 2020;Sue et al, 2023). On the other hand, the adhesive tip pilin, which has the sorting motif but lacks the linking lysine or pilin motif, can only be added to the top of the pilus structure, presumably at the start of the polymerization process (Krishnan et al, 2013;Izore ´et al, 2010;Kant et al, 2020;Yadav & Krishnan, 2022).…”