A specific, Na+-dependent, energy-requiring transport system for taurine has been reported recently in the Staphylococcus aureus M strain. Taurine was taken up vigorously by all S . aureus strains tested. The system was Na+-dependent, and Na+ decreased the K , but had no effect on the V,,, of the transport system. Among coagulase-negative staphylococci, the Staphylococcus epidermidis group (a taxonomically related group of species associated with humans or other primates) and the free-living, wide-ranging species Staphylococcus sciuri showed vigorous taurine uptake. Somewhat lower rates were found in the Staphylococcus saprophyticus group. Low or barely detectable uptake rates were noted in other staphylococcal species that were primarily of animal origin. N o taurine uptake was detected in a variety of other bacterial species tested. Taurine uptake, which was not Na+-dependent, occurred in a Pseudomonas aeruginosa strain grown on taurine as sole energy, carbon, nitrogen, and sulphur source, but not when it was grown in a gluconate/salts medium. In nutritional studies we were unable to demonstrate a role for taurine as a sulphur source for S . aureus. [ 1 ,2-14C]-and [35S]taurine were taken up during overnight growth of cells, and radioactivity was distributed similarly among cellular fractions, indicating that the carbon and sulphur atoms of taurine were not cleaved and had the same fate.We were unable to demonstrate any catabolism of taurine in radiorespirometric experiments to detect evolution of I4CO2 by cells incubated with [ 1 ,2-14C]taurine. Thus, we found no evidence for a role of taurine in the energy, carbon and sulphur metabolism of S . aureus.
Hemolytic-Uremic Syndrome (HUS) clinically presents as a triad of hemolytic anemia, thrombocytopenia and acute renal failure. Mostly, it is caused by infection with certain strains of Shiga toxin-producing E. coli. Recently, a novel class of single-domain antibodies (VHH) or “nanobodies” was raised against Shiga toxin (Stx) and was shown to protect Stx-exposed mice. However, Stx is cleared rapidly from the circulation and is generally not detectable at the time of systemic disease onset. We investigated whether anti-Stx nanobodies (Nbs) can be effective in neutralizing Stx intracellularly. Two Nbs with high affinity to the B subunit of Stx2a were selected to investigate their effect against Shiga toxin. The Nb sequence was cloned in-frame into a backbone vector carrying an Endoplasmic Reticulum (ER) localization and retention signals (KDEL), and Green Fluorescence Protein (GFP). Shiga toxin-sensitive cells (hCMEC/D3, and BxPc3) were transfected with the vector, a cytotoxic dose of the toxin was added after 24 hours and cells viability were determined by resazurin assay at different time points. Intracellular ER-localized expression of anti-Stx nanobodies (JFL47, JGL-34) also the ER-backbone vectors without nanobody insert protected the cell lines at all timepoints. Control plasmids of other signaling peptides [mem: signal sequence targeting plasma membrane+GFP as positive control] or unrelated vectors [N1: empty vector, GFP alone as negative control] did not show significant differences to wildtype. While inconclusive regarding the protective effect of anti-Shiga toxin nanobody constructs, a novel and significant finding of this study is the observed intrinsic, highly protective effect of the ERtargeting sequence (SEKDEL/KDEL).
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