4 years ago

Structural and functional dissection reveals distinct roles of Ca<sup>2+</sup>-binding sites in the giant adhesin SiiE of <i>Salmonella enterica</i>

Johanna Stein, Achim Sandmann, Britta Peters, Stefan Klingl, Heinrich Sticht, Yves A. Muller, Nicola Taccardi, Michael Hensel, Roman G. Gerlach, Nathalie Sander

by Britta Peters, Johanna Stein, Stefan Klingl, Nathalie Sander, Achim Sandmann, Nicola Taccardi, Heinrich Sticht, Roman G. Gerlach, Yves A. Muller, Michael Hensel

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.

Publisher URL: http://journals.plos.org/plosone/article

DOI: 10.1371/journal.ppat.1006418

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