X-ray crystallographic studies of streptavidin mutants binding to biotin

On the basis of high resolution crystallographic studies ot streptavidin an d its biotin complex, three principal binding motifs have been identified t hat contribute to the tight binding. A flexible binding loop can undergo a conformational change from an open to a closed form when biotin is bound. A dditional studies described here of unbound wild-type streptavidin have pro vided structural views of the open conformation. Several tryptophan residue s packing around the bound biotin constitute the second binding motif, one dominated by hydrophobic interactions. Mutation of these residues to alanin e or phenylalanine have variable effects on the thermodynamics and kinetics of binding, but they generate only small changes in the molecular structur e. Hydrogen bonding interactions also contribute significantly to the bindi ng energetics of biotin, and the D128A mutation which breaks a hydrogen bon d between the protein and a ureido NH group results in a significant struct ural alteration that could mimic an intermediate on the dissociation pathwa y. In this review, we: summarize the structural aspects of biotin recogniti on that have been gained from crystallographic analyses of wild-type and si te-directed streptavidin mutants. (C) 1999 Published by Elsevier Science B. V. All rights reserved.