The quaternary structure of streptavidin in urea

• Published in: The Journal of biological chemistry 1991-08, Vol.266 (22), p.14470-14477
• Main Authors: KURZBAN, G. P, BAYER, E. A, WILCHEK, M, HOROWITZ, P. M
• Format: Article
• Language: English
• Subjects: Analytical, structural and metabolic biochemistry; avidin; Bacterial Proteins - chemistry; Biological and medical sciences; biotin; Biotin - chemistry; Chromatography, Gel; Electrophoresis, Polyacrylamide Gel; Fundamental and applied biological sciences. Psychology; Guanidine; Guanidines - chemistry; Miscellaneous; Protein Conformation; Proteins; Spectrometry, Fluorescence; Streptavidin; urea; Urea - chemistry
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/s0021-9258(18)98710-7

We report on the interactions of urea and guanidinium salts with streptavidin. Gel filtration chromatography in 0, 4, 6, and 7 M urea indicates that the streptavidin tetramer remains intact in urea. Biotin alters the electrophoretic mobility of streptavidin whether or not 6 M urea is present. The intrinsic fluorescence of streptavidin is increased and blue-shifted in 6 M urea. The fluorescence changes indicate the absence of unfolding. A conformational response to urea is possible, but much of the fluorescence change is due to urea binding as a weak biotin analog (Ka approximately 1.3 M-1). The resistance to structural perturbation by urea reflects the structural stability of streptavidin's anti-parallel beta-barrel motif. Unfolding is sluggish in 6 M guanidinium hydrochloride (half-time, approximately 50 days). After guanidinium thiocyanate unfolding, streptavidin can be refolded, but the unfolding and refolding transitions are centered at different concentrations of perturbant. Slow unfolding, with a 15th power dependence on guanidinium thiocyanate concentration, may be partially responsible for the noncoincidence of the unfolding and refolding processes. Nonequilibrium behavior is also seen in 6 M urea, as native streptavidin does not unfold and guanidinium thiocyanate unfolded streptavidin does not refold. Refolding does occur at lower concentrations of urea. Guanidinium thiocyanate only slowly unfolds the biotin-streptavidin complex. In the presence of biotin, unfolded streptavidin does not refold in 6 M guanidinium thiocyanate or in 6 M urea.