Iron-regulated surface determinant B (IsdB) promotes Staphylococcus aureus adherence to and internalization by non-phagocytic human cells
Article first published online: 15 JAN 2013
© 2012 John Wiley & Sons Ltd
Volume 15, Issue 6, pages 1026–1041, June 2013
How to Cite
Zapotoczna, M., Jevnikar, Z., Miajlovic, H., Kos, J. and Foster, T. J. (2013), Iron-regulated surface determinant B (IsdB) promotes Staphylococcus aureus adherence to and internalization by non-phagocytic human cells. Cellular Microbiology, 15: 1026–1041. doi: 10.1111/cmi.12097
- Issue published online: 14 MAY 2013
- Article first published online: 15 JAN 2013
- Accepted manuscript online: 24 DEC 2012 02:45AM EST
- Manuscript Accepted: 13 DEC 2012
- Manuscript Revised: 28 NOV 2012
- Manuscript Received: 30 JUL 2012
- Health Research Board of Ireland. Grant Number: RP/2008/20
- Science Foundation Ireland Principle Investigator. Grant Number: 08/IN.1/B1854
- Slovenian Research Agency. Grant Number: J4-0123
Fig. S1. Recombinant IsdB and IsdB Y165A/H166E/Y167A binding to human haemoglobin (hHb). rIsdB was immobilized in and incubated with different concentration of hHb. Bound hHb was detected with polyclonal rabbit anti-hHb serum followed by HRP-conjugated goat anti-rabbit IgG. Values represent the mean of triplicate wells. Binding assays was preformed three times with similar results.
Fig. S2. Inducible expression of IsdB by L. lactis. Whole-cell immunoblot to detect surface expression of IsdB by L. lactis NZ9000. L. lactis pNZ8037isdB and L. lactis pNZ8037 were grown to exponential phase and induced with nisin (at indicated concentrations). After 4 h post induction the bacteria were harvested, dotted on cellulose membranes (at indicted concentrations) and subjected to immunoblotting. Detection was performed using rabbit anti-IsdB IgG followed by HRP-conjugated goat anti-rabbit IgG.
Fig. S3. Real-time measurement of attachment and spreading of breast cancer MCF-7 cells. (A) Fibronectin (Fn) was immobilized in microtitre wells, blocked with BSA and incubated with equal numbers of MCF-7 cells supplemented with either IsdA, IsdH-GST, GST or IsdB. XCELLigence was used to measure the real-time impedance in those wells. Cells attachment and spreading led to increased impedance which corresponded to higher cell index (CI) values. (A) Traces of CI changes representing MCF-7 spreading on Fn in the presence of IsdA (light grey), IsdH-GST (middle grey), GST (dark grey) or IsdB (black) over time are shown. (B) Slope changes (slope h−1) of CI values representing MCF-7 cells attachment to fibronectin upon incubation with different proteins (as indicated). Slope changes were calculated for CI traces within time points of 30 min to 240 min.
Table S1. Primers used in this study.
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