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Cooperative hydrogen bond interactions in the streptavidin–biotin system
Author(s) -
Hyre David E.,
Le Trong Isolde,
Merritt Ethan A.,
Eccleston John F.,
Green N. Michael,
Stenkamp Ronald E.,
Stayton Patrick S.
Publication year - 2006
Publication title -
protein science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.353
H-Index - 175
eISSN - 1469-896X
pISSN - 0961-8368
DOI - 10.1110/ps.051970306
Subject(s) - hydrogen bond , chemistry , biotin , crystallography , cooperativity , mutant , enthalpy , streptavidin , binding energy , cooperative binding , stereochemistry , binding site , molecule , biochemistry , thermodynamics , physics , atomic physics , organic chemistry , gene
The thermodynamic and structural cooperativity between the Ser45– and D128–biotin hydrogen bonds was measured by calorimetric and X‐ray crystallographic studies of the S45A/D128A double mutant of streptavidin. The double mutant exhibits a binding affinity ∼2 × 10 7 times lower than that of wild‐type streptavidin at 25°C. The corresponding reduction in binding free energy (ΔΔ G ) of 10.1 kcal/mol was nearly completely due to binding enthalpy losses at this temperature. The loss of binding affinity is 11‐fold greater than that predicted by a linear combination of the single‐mutant energetic perturbations (8.7 kcal/mol), indicating that these two mutations interact cooperatively. Crystallographic characterization of the double mutant and comparison with the two single mutant structures suggest that structural rearrangements at the S45 position, when the D128 carboxylate is removed, mask the true energetic contribution of the D128–biotin interaction. Taken together, the thermodynamic and structural analyses support the conclusion that the wild‐type hydrogen bond between D128–OD and biotin–N2 is thermodynamically stronger than that between S45–OG and biotin–N1.