Structural and functional dissection reveals distinct roles of Ca2+-binding sites in the giant adhesin SiiE of Salmonella enterica

PLoS Pathog. 2017 May 30;13(5):e1006418. doi: 10.1371/journal.ppat.1006418. eCollection 2017 May.


The giant non-fimbrial adhesin SiiE of Salmonella enterica mediates the first contact to the apical site of epithelial cells and enables subsequent invasion. SiiE is a 595 kDa protein composed of 53 repetitive bacterial immunoglobulin (BIg) domains and the only known substrate of the SPI4-encoded type 1 secretion system (T1SS). The crystal structure of BIg50-52 of SiiE revealed two distinct Ca2+-binding sites per BIg domain formed by conserved aspartate or glutamate residues. In a mutational analysis Ca2+-binding sites were disrupted by aspartate to serine exchange at various positions in the BIg domains of SiiE. Amounts of secreted SiiE diminish with a decreasing number of intact Ca2+-binding sites. BIg domains of SiiE contain distinct Ca2+-binding sites, with type I sites being similar to other T1SS-secreted proteins and type II sites newly identified in SiiE. We functionally and structurally dissected the roles of type I and type II Ca2+-binding sites in SiiE, as well as the importance of Ca2+-binding sites in various positions of SiiE. Type I Ca2+-binding sites were critical for efficient secretion of SiiE and a decreasing number of type I sites correlated with reduced secretion. Type II sites were less important for secretion, stability and surface expression of SiiE, however integrity of type II sites in the C-terminal portion was required for the function of SiiE in mediating adhesion and invasion.

MeSH terms

  • Adhesins, Bacterial / chemistry*
  • Adhesins, Bacterial / genetics
  • Adhesins, Bacterial / metabolism*
  • Amino Acid Motifs
  • Bacterial Adhesion
  • Bacterial Proteins / chemistry*
  • Bacterial Proteins / genetics
  • Bacterial Proteins / metabolism*
  • Calcium / metabolism*
  • Salmonella Infections / microbiology*
  • Salmonella typhimurium / chemistry
  • Salmonella typhimurium / genetics
  • Salmonella typhimurium / physiology*


  • Adhesins, Bacterial
  • Bacterial Proteins
  • Calcium

Grant support

This work was funded by the Deutsche Forschungsgemeinsschaft ( though grants: SFB944/P4 to MH, HE1964/13-2 to MH, MU1377/9-2 to YAM, GE2533/2-2 to RGG, SFB796 A3 to YAM, and SFB796 A2 to HS. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.