A research team led by the University of Illinois has developed a treatment for exposure to enterotoxin B, a noxious substance produced by the Staphylococcus aureus bacterium. The team engineered a protein, which was successfully tested in rabbits, that could one day be used to treat humans exposed to the enterotoxin.
S. aureus enterotoxin B (SEB) is a common cause of food poisoning, but if it is inhaled or produced during an infection it can elicit a systemic -- and sometimes fatal -- immune response in humans. In purified form, SEB is listed as a potential bioterrorism agent. Other potent S. aureus enterotoxins include the toxic shock syndrome toxin.
These enterotoxins are classed as superantigens because they set off a massive immune response in humans and other animals. They bind to variable regions of T-cell receptors, stimulating a cascade of events, including the systemic release of inflammatory cytokines. In some cases the powerful immune response leads to toxic shock and death.
The research team, led by University of Illinois professor of biochemistry David M. Kranz, included scientists and clinicians from the Boston Biomedical Research Institute and the University of Minnesota Medical School.
The team began by engineering a protein with the same structure as the binding site of the T-cell receptor targeted by SEB. The researchers expressed the engineered protein on the surface of yeast cells (using a process they helped develop, called "yeast display") and generated mutations meant to increase the protein's ability to bind SEB. After several rounds of mutagenesis and screening, graduate student Rebecca A. Buonpane developed a soluble protein with an affinity for SEB that was over a million times that of the original.
"Our approach was to take these receptors that bind to the toxins and to try to make them higher affinity and therefore act as effective neutralizing agents when delivered in soluble form," Kranz said. "It's the binding of the toxin to T-cells that is critical. If you can prevent the toxin from binding to the T-cell receptor then you can prevent it from initiating that cascade."
The engineered protein prevented the onset of symptoms in rabbits exposed to SEB and reversed the course of the illness in those treated two hours after exposure. "We were very pleasantly surprised that it showed effectiveness in every rabbit tested," Kranz said.
He noted that the protein has some potential advantages and disadvantages when compared to antibodies, which might also be used to fight infection with SEB. One advantage is that the engineered protein is small, about one-tenth the size of an antibody. Its size may allow it to penetrate deeper into tissues, and may make it less likely to spark an immune response in animals. The protein can also be produced in large quantities using the bacterium, Escherichia coli.
"E. coli is the cheapest source for making proteins," Kranz said. "Whenever you can express a protein in E. coli you do so because it is inexpensive, easy and fast."
Antibodies, on the other hand, can remain in the body for days or weeks, whereas the new protein is cleared within hours. This may make antibodies a better treatment option in some circumstances, said Kranz.
No antibody has yet been developed, however, that has a comparable affinity for SEB.
The findings appear in the online edition of Nature Medicine. These studies were supported by the National Institute of Allergy and Infectious Diseases, part of the National Institutes of Health.
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