Abstract
The human pathogen Streptococcus pyogenes produces diverse pili depending on the serotype. We investigated the assembly mechanism of FCT type 1 pili in a serotype M6 strain. The pili were found to be assembled from two precursor proteins, the backbone protein T6 and ancillary protein FctX, and anchored to the cell wall in a manner that requires both a housekeeping sortase enzyme (SrtA) and pilus-associated sortase enzyme (SrtB). SrtB is primarily required for efficient formation of the T6 and FctX complex and subsequent polymerization of T6, whereas proper anchoring of the pili to the cell wall is mainly mediated by SrtA. Because motifs essential for polymerization of pilus backbone proteins in other Gram-positive bacteria are not present in T6, we sought to identify the functional residues involved in this process. Our results showed that T6 encompasses the novel VAKS pilin motif conserved in streptococcal T6 homologues and that the lysine residue (Lys-175) within the motif and cell wall sorting signal of T6 are prerequisites for isopeptide linkage of T6 molecules. Because Lys-175 and the cell wall sorting signal of FctX are indispensable for substantial incorporation of FctX into the T6 pilus shaft, FctX is suggested to be located at the pilus tip, which was also implied by immunogold electron microscopy findings. Thus, the elaborate assembly of FCT type 1 pili is potentially organized by sortase-mediated cross-linking between sorting signals and the amino group of Lys-175 positioned in the VAKS motif of T6, thereby displaying T6 and FctX in a temporospatial manner.
Highlights
The variability of S. pyogenes pili on both the gene and protein levels is outstanding among streptococcal species, which is reflected by the fact that the antigenicity of pilus proteins is one of the determinants for T serotyping [20, 23, 24, 41]
To begin molecular characterization of far uncharacterized pili, we focused on the assembly mechanism of FCT type 1 pili in a serotype M6 strain
In a serotype M6 strain, two sortase enzymes are encoded in the genome, i.e. the housekeeping sortase SrtA and pilus region-associated sortase SrtB [8]
Summary
Polymerization of the major subunit constitutes the pilus backbone shaft, whereas the minor subunits are covalently incorporated into the backbone as ancillary pilus proteins These pilus subunits contain cell wall sorting signals (CWSSs), including a canonical LPXTG or LPXTG-like motif [5]. Pilus subunits are assembled via this motif, i.e. polymerization of major subunits and formation of major and minor subunit complexes, and cross-linked to the peptidoglycan layer These processes are mediated by a membrane-bound transpeptidase termed sortase, including a housekeeping sortase enzyme (SrtA) and pilus-associated sortase enzyme (SrtB in this study) (6 – 8). We found a lysine residue within a novel VAKS pilin motif of T6 required for T6 polymerization and incorporation of FctX at the tip of the pilus shaft These unique characteristics of the assembly mechanism of FCT type 1 pili give insights into the complexity of the biological nature of streptococcal pili
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
