4L6G

Crystal Structure of P450cin Y81F mutant, crystallized in 7 mM 1,8-cineole

4L6G の概要

• エントリーDOI: 10.2210/pdb4l6g/pdb
• 関連するPDBエントリー: 4L77 4LHT
• 分子名称: P450cin, PROTOPORPHYRIN IX CONTAINING FE, 1,3,3-TRIMETHYL-2-OXABICYCLO[2.2.2]OCTANE, ... (6 entities in total)
• 機能のキーワード: p450, heme, monooxygenase, cindoxin, oxidoreductase
• 由来する生物種: Citrobacter braakii
• タンパク質・核酸の鎖数: 2
• 化学式量合計: 91673.77

構造登録者

Madrona, Y.,Poulos, T.L. (登録日: 2013-06-12, 公開日: 2013-07-24, 最終更新日: 2023-09-20)

主引用文献

Madrona, Y.,Hollingsworth, S.A.,Khan, B.,Poulos, T.L.
P450cin active site water: implications for substrate binding and solvent accessibility.
Biochemistry, 52:5039-5050, 2013

PubMed Abstract: In P450cin, Tyr81, Asp241, Asn242, two water molecules, and the substrate participate in a complex H-bonded network. The role of this H-bonded network in substrate binding and catalysis has been probed by crystallography, spectroscopy, kinetics, isothermal titration calorimetry (ITC), and molecular dynamics. For the Y81F mutant, the substrate binds about 20-fold more weakly and Vmax decreases by about 30% in comparison to WT. The enhanced susceptibility of the heme to H₂O₂-mediated destruction in Y81F suggests that this mutant favors the open, low-spin conformational state. Asn242 H-bonds directly with the substrate, and replacing this residue with Ala results in water taking the place of the missing Asn side chain. This mutant exhibits a 70% decrease in activity. Crystal structures and molecular dynamics simulations of substrate-bound complexes show that the solvent has more ready access to the active site, especially for the N242A mutant. This accounts for about a 64% uncoupling of electron transfer from substrate hydroxylation. These data indicate the importance of the interconnected water network on substrate binding and on the open/closed conformational equilibrium, which are both critically important for maintaining high-coupling efficiency.

PubMed: 23829586
DOI: 10.1021/bi4006946

実験手法

X-RAY DIFFRACTION (1.371 Å)