The binding of biotin analogues by streptavidin: A Raman spectroscopic study

Abstract

Raman spectra of anhydrous complexes of streptavidin (Strep) with biotin (Bio) and some Bio analogues [Biotin methyl ester (MEBio), desthiobiotin (DEBio), 2′-iminobiotin (IMBio), and diaminobiotin (DABio)] were recorded. The vibrational results indicate that the interaction with some of these ligands is able to modify the overall structure of the protein and this binding results in a decrease in the βsheet content and an increase in the α-helix content. To further confirm the conformational changes of the protein structure due to Bio analogue binding, the curve-fitting analysis of the amide I Raman band of neat Strep and of the complexes were performed. The intensity ratio of the components due to the β-sheet and α-helix conformations decreased in the Strep-MEBio, Strep-IMBio (pH 11), and Strep-Bio systems, whereas in all the other systems the changes were not significant. This behavior differs from that of Avi bound to the same ligands and suggests that Strep and Avi differ in their binding selectivity. A good correlation was found between the secondary structure percentages of the Avi and of the Strep complexes and ΔG°. On the basis of this linear relationship, the vibrational results allow for an acceptable evaluation of the dissociation constants of the Strep complexes, not previously reported in the literature. The present results indicate a correlation between the type of interaction and the effects of the protein-substrate bonding on the overall structure of the proteins. The amino acid residues in the binding site appear to be positioned in a such a way as to provide a precise fit of Bio. Even slight change in the substrate structure causes a weakness in the strength of the binding. The vibrational results confirm that both the imidazolidinone and the thiophan rings are important in the Strep-Bio interactions, but the former is more responsible for the high affinity of the binding. One of the Tyr residues is hydrogen bound with the ureido ring and another Tyr could be involved in the binding pocket. Trp residues do not directly bind the ligand and probably stabilize other binding site residues which in turn interact directly with Bio. © 1998 John Wiley & Sons, Inc. Biospectroscopy 4: 197–208, 1998