1df8

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spinqualitylabelsall models 1df8, resolution 1.51Å Show:Asymmetric Unit Biological Assembly Drag the structure with the mouse to rotate   Export Animated Image

S45A MUTANT OF STREPTAVIDIN IN COMPLEX WITH BIOTIN

Overview

The contribution of the Ser45 hydrogen bond to biotin binding activation, and equilibrium thermodynamics was investigated by biophysical and X-ray, crystallographic studies. The S45A mutant exhibits a 1,700-fold greater, dissociation rate and 907-fold lower equilibrium affinity for biotin, relative to wild-type streptavidin at 37 degrees C, indicating a crucial, role in binding energetics. The crystal structure of the biotin-bound, mutant reveals only small changes from the wild-type bound structure, and, the remaining hydrogen bonds to biotin retain approximately the same, lengths. No additional water molecules are observed to replace the missing, hydroxyl, in contrast to the previously studied D128A mutant. The, equilibrium deltaG degrees, deltaH degrees, deltaS degrees, deltaC, degrees(p), and activation deltaG++ of S45A at 37 degrees C are 13.7+/-0.1, kcal/mol, -21.1+/-0.5 kcal/mol, -23.7+/-1.8 cal/mol K, -223+/-12 cal/mol, K, and 20.0+/-2.5 kcal/mol, respectively. Eyring analysis of the large, temperature dependence of the S45A off-rate resolves the deltaH++ and, deltaS++ of dissociation, 25.8+/-1.2 kcal/mol and 18.7+/-4.3 cal/mol K., The large increases of deltaH++ and deltaS++ in the mutant, relative to, wild-type, indicate that Ser45 could form a hydrogen bond with biotin in, the wild-type dissociation transition state, enthalpically stabilizing it, and constraining the transition state entropically. The postulated, existence of a Ser45-mediated hydrogen bond in the wild-type streptavidin, transition state is consistent with potential of mean force simulations of, the dissociation pathway and with molecular dynamics simulations of biotin, pullout, where Ser45 is seen to form a hydrogen bond with the ureido, oxygen as biotin slips past this residue after breaking the native, hydrogen bonds.

About this Structure

1DF8 is a Single protein structure of sequence from Streptomyces avidinii with BTN as ligand. Full crystallographic information is available from OCA.

Reference

Ser45 plays an important role in managing both the equilibrium and transition state energetics of the streptavidin-biotin system., Hyre DE, Le Trong I, Freitag S, Stenkamp RE, Stayton PS, Protein Sci. 2000 May;9(5):878-85. PMID:10850797

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