Peptide ligands that bind to staphylococcal enterotoxin B (SEB)

Staphylococcal enterotoxin B (SEB) is a primary toxin that causes food poisoning. It also acts as a superantigen that interacts with the major histocompatibility complex class II molecule (MHCII) and T cell receptor (TCR) to activate large amounts of T cells leading to autoimmune diseases. Highly purified SEB is needed in research and can be used as a standard in current detection methods for SEB. SEB is also a contaminant in protein A purification from Staphylococcal aureus fermentation broth. Inexpensive, robust ligands with high affinity would be suitable replacements for antibodies in biosensors for the detection of SEB. Affinity adsorption processes using short peptides as ligands show great promise in purifying and detecting proteins in comparison with other methods due to their high stability and low cost. By screening a solid-phase combinatorial peptide library, a short peptide ligand, YYWLHH, has been discovered that binds with high affinity and selectivity to SEB, but only weakly to other staphylococcal enterotoxins that share sequence and structural homology with SEB. Using column affinity chromatography with an immobilized YYWLHH stationary phase, it was possible to separate SEB quantitatively from Staphylococcus aureus fermentation broth, a complex mixture of proteins, carbohydrates and other biomolecules. The immobilized peptide was also used to purify native SEB from a mixture containing denatured and hydrolyzed SEB, and showed little cross reactivity with other SEs. To our knowledge this is the first report of a highly specific short peptide ligand for SEB. Such a ligand is a potential candidate to replace antibodies for detection, removal and purification strategies for SEB.Modeling the transport and kinetic processes in peptide affinity chromatography allows for a direct measurement of the rate of binding of SEB to peptide ligands. It can also provide design parameters for columns that can be used to either remove or detect SEB as well as other pathogens. The mass transfer parameters of SEB were either measured using pulse experiments or determined from correlations. The adsorption isotherms of SEB on YYWLHH resins with different peptide densities were fitted to bi-Langmuir isotherms. The general rate (GR) model was used to fit experimental breakthrough curves to obtain the intrinsic rate constants of the adsorption-desorption kinetics of the protein on the peptide ligands. An analysis of the number of transfer units in the column revealed that both intraparticle mass transfer and intrinsic adsorption rates were important rate-limiting steps for adsorption to the resin particles. (Abstract shortened by UMI.)...