Many pathogenic bacteria express proteins that interfere with the host defense, with complement evasion a central strategy to their success in causing infection and disease 
. As a highly successful pathogen, S. aureus
is no exception. S. aureus
produces an effective arsenal directly targeted at the host immune system, including the complement cascade and inhibition of its downstream effects. Our current knowledge of the ability of S. aureus
to subvert the complement system continues to grow.
In the present study, we identified two potential fH-binding S. aureus
cell wall proteins: SdrE and ClfA. Both belong to the Sdr family of structurally related cell wall-associated proteins that contain a region of serine-aspartate repeats 
. Anchored via a conserved LPXTG motif, SdrE and ClfA possess an R region containing the repeating SD dipeptides, and a unique A region; SdrE also contains a B region. ClfA is known to bind fibrinogen 
and fI, as well as function as a cofactor for fI-mediated degradation of C3b 
. SdrE is less well described; however, it is implicated in human platelet aggregation when expressed on the surface of Lactococcus lactis
. Until now, a definitive ligand for SdrE has been elusive.
We identified SdrE as a putative fH-binding protein via mass spectrometric analysis of S. aureus
cell wall proteins isolated by two distinct techniques: purified fH overlay blot of fractionated cell wall proteins as well as cross-linking. The corresponding identifications provided extremely high peptide scores and expect values indicating the strength of these identifications. Studies using recombinantly expressed SdrE validated the ability of this protein to bind fH whether purified or in serum, with a time- and dose-dependent relationship evident. Using a gain-of-function L. lactis
model, we demonstrated that SdrE expression on a bacterial surface significantly enhances fH recruitment, which confirms our rSdrE-fH binding data. Functional analysis of rSdrE-bound fH revealed that fH remains functionally active in its ability to provide cofactor activity for fI-mediated cleavage of C3b, with a positive correlation of iC3b generation for increasing amounts of fH. This was also observed for L. lactis
-SdrE-bound fH. Cleaved C3b (iC3b) can no longer participate in the formation of C3- and C5-convertases which negatively affects amplification of the complement cascade 
resulting in decreased S. aureus
phagocytosis, as we have previously demonstrated 
. Likewise, surface expression of SdrE on the surrogate bacterium L. lactis
resulted in less C3-fragment deposition, less C5a generation, and decreased complement-mediated killing by neutrophils. The down-regulation of complement-mediated host defenses in this gain-of-function model strongly suggests that SdrE is an immune evasion protein. Indeed, S. aureus
strains that express SdrE are typically associated with invasive infection 
with 90% of 497 S. aureus
isolates tested being sdrE
While characterizing the fH-binding protein band fractionated from cell wall preparations, we also identified ClfA. However, no ClfA peptides were identified in the cross-linked sample. Recombinantly expressed ClfA did not bind purified fH and anti-fH Western-blot analysis of serum proteins bound to rClfA revealed no significant difference between fH binding to rClfA compared to BSA, as determined via optical densitometry. Therefore, it seems reasonable that ClfA was originally identified along with SdrE as a putative fH-binding protein due to co-migration during fractionation and gel electrophoresis attributable to the similar molecular weights and charges of these proteins.
Our earlier examination of S. aureus
-bound serum proteins suggests that S. aureus
binds FHL-1, FHR-1α, and/or FHR-1β 
. Therefore, it is possible that these proteins interact with SdrE. However, under the conditions tested, we were unable to demonstrate binding of FHL-1, FHR-1α, and/or FHR-1β to recombinant SdrE or SdrE expressed on the surface of L. lactis
. The serum concentration of FHL-1 is 10–50 times lower than fH 
, which may have contributed to the negative result. FHR-1α and FHR-1β are known to be at a much lower concentration in serum than fH; however, their concentrations have not been clearly defined 
. Therefore, whether FHL-1 and/or FHR-1α/β interact with SdrE will be further addressed in future studies.
The staphylococcus protein Sbi has previously been shown to bind fH in a triparte complex with C3b via Sbi domains III and IV 
. Sbi can be found in both the cytoplasmic membrane fraction as well as secreted into the external milieu 
. The N-terminal domains of Sbi (I and II) bind IgG in a similar manner to staphylococcal protein A when exposed on the cell surface, whereas the C-terminal domains of Sbi (III and IV) are only biologically active when secreted 
. Therefore, Sbi cannot contribute to the acquisition of fH to the staphylococcal surface.
In summary, our data show that rSdrE is a clear fH-binding molecule due to its ability to bind fH whether purified or in serum. Moreover, rSdrE-bound fH retains cofactor activity for fI-mediated cleavage of C3b. Additionally, our studies using SdrE-expressing L. lactis demonstrate that SdrE expression on the bacterial surface increases the binding of fH, down-regulates complement effectors, and provides protection from neutrophil killing. As such, SdrE recruitment of fH likely provides a survival advantage for S. aureus by negatively affecting the formation of complement activating complexes, thereby dampening the host immune response. To our knowledge, this is the first description of a S. aureus surface protein that recruits the potent complement regulator fH to evade the immune response.