The spectral and thermodynamic properties of staphylococcal enterotoxin A, E, and variants suggest that structural modifications are important to control their function.
The superantigens staphylococcal enterotoxin A and E (SEA and SEE) can activate a large number of T-cells. SEA and SEE have approximately 80% sequence identity but show some differences in their biological function. Here, the two superantigens and analogues were characterized biophysically. SEE was shown to have a substantially higher thermal stability than SEA. Both SEA and SEE were thermally stabilized by 0.1 mM Zn(2+) compared with Zn(2+)-reduced conditions achieved using 1 mM EDTA or specific replacements that affect Zn(2+) coordination. The higher stability of SEE was only partly caused by the T-cell receptor (TCR) binding regions, whereas regions in the vicinity of the major histocompatibility complex class II binding sites affected the stability to a greater extent. SEE exhibited a biphasic denaturation between pH 5.0-6.5, influenced by residues in the TCR binding regions. Interestingly, enzyme-linked immunosorbent assay, isoelectric focusing, and circular dichroism analysis indicated that conformational changes had occurred in the SEA/E chimerical constructs relative to SEA and SEE. Thus, it is proposed that the Zn(2+) binding site is very important for the stability and potency of SEA and SEE, whereas residues in the TCR binding site have a substantial influence on the molecular conformation to control specificity and function.
Cavallin
A
A
Active Biotech Research AB, Box 724, 22007 Lund, Sweden.
Arozenius
H
H
Kristensson
K
K
Antonsson
P
P
Otzen
D E
DE
Björk
P
P
Forsberg
G
G
D000595:Amino Acid Sequence; D000818:Animals; D002478:Cells, Cultured; D002942:Circular Dichroism; D004305:Dose-Response Relationship, Drug; D004768:Enterotoxins / Q000737:chemistry*; D019791:Guanidine / Q000494:pharmacology; D000949:Histocompatibility Antigens Class II / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D051379:Mice; D008810:Mice, Inbred C57BL; D008969:Molecular Sequence Data; D011485:Protein Binding; D011489:Protein Denaturation; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry; D017386:Sequence Homology, Amino Acid; D013154:Spleen / Q000378:metabolism; D013329:Structure-Activity Relationship; D018089:Superantigens / Q000378:metabolism; D013601:T-Lymphocytes / Q000378:metabolism; D013696:Temperature; D013816:Thermodynamics; D015032:Zinc / Q000378:metabolism
D004768:Enterotoxins; D000949:Histocompatibility Antigens Class II; D011993:Recombinant Fusion Proteins; D018089:Superantigens; C049463:enterotoxin E, Staphylococcal; C026066:enterotoxin A, Staphylococcal; D015032:Zinc; D019791:Guanidine