Role of glycosylation at Ser63 in production of soluble pilin in pathogenic Neisseria

Pilus-mediated adhesion is essential in the pathogenesis of Neisseria menin gitidis (MC) and Neisseria gonorrhoeae (GC). Pili are assembled from a prot ein subunit called pilin. Pilin is a glycoprotein, and pilin antigenic vari ation has been shown to be responsible for intrastrain variability with res pect to the degree of adhesion in both MC and GC. In MC, high-adhesion pili ns are responsible for the formation of bundles of pill which bind bacteria and cause them to grow as colonies on infected monolayers. In this work, w e selected MC and GC pilin variants responsible for high and low adhesivene ss and introduced them into the other species. Our results demonstrated tha t a given pilin variant expressed an identical phenotype in either GC or MC with respect to bundling and adhesiveness to epithelial cells. However, th e production of truncated soluble pilin (S pilin) was consistently more abu ndant in GC than in MC. In the latter species, the glycosylation of pilin a t Ser63 was shown to be required for the production of a truncated monomer of S pilin. In order to determine whether the same was true for GC, we engi neered various pilin derivatives with an altered Ser63 glycosylation site. The results of these experiments demonstrated that the production of S pili n in GC was indeed more abundant when pilin was posttranslationally modifie d at Ser63. However, nonglycosylated variants remained capable of producing large amounts of S pilin. These data demonstrated that for GC, unlike for MC, glycosylation at Ser63 is not required for S-pilin production, suggesti ng that the mechanisms leading to the production of S pilin in GC and MC ar e different.