1DF8

S45A MUTANT OF STREPTAVIDIN IN COMPLEX WITH BIOTIN

1DF8 の概要

• エントリーDOI: 10.2210/pdb1df8/pdb
• 分子名称: PROTEIN (STREPTAVIDIN), BIOTIN (3 entities in total)
• 機能のキーワード: biotin binding protein, biotin, binding protein
• 由来する生物種: Streptomyces avidinii
• 細胞内の位置: Secreted: P22629
• タンパク質・核酸の鎖数: 2
• 化学式量合計: 27019.29

構造登録者

Hyre, D.E.,Le Trong, I.,Freitag, S.,Stenkamp, R.E.,Stayton, P.S. (登録日: 1999-11-18, 公開日: 2000-09-20, 最終更新日: 2023-08-09)

主引用文献

Hyre, D.E.,Le Trong, I.,Freitag, S.,Stenkamp, R.E.,Stayton, P.S.
Ser45 plays an important role in managing both the equilibrium and transition state energetics of the streptavidin-biotin system.
Protein Sci., 9:878-885, 2000

PubMed Abstract: 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.

PubMed: 10850797

実験手法

X-RAY DIFFRACTION (1.51 Å)