5KP4

Crystal Structure of Ketosteroid Isomerase from Pseudomonas putida (pKSI) bound to 19-nortestosterone

5KP4 の概要

• エントリーDOI: 10.2210/pdb5kp4/pdb
• 関連するPDBエントリー: 5D81 5D82 5KP1 5KP3
• 分子名称: Steroid Delta-isomerase, (8~{R},9~{S},10~{R},13~{S},14~{S},17~{S})-13-methyl-17-oxidanyl-2,6,7,8,9,10,11,12,14,15,16,17-dodecahydro-1~{H}-cyclop enta[a]phenanthren-3-one (3 entities in total)
• 機能のキーワード: isomerase
• 由来する生物種: Pseudomonas putida
• タンパク質・核酸の鎖数: 2
• 化学式量合計: 30304.60

構造登録者

Wu, Y.,Boxer, S.G. (登録日: 2016-07-01, 公開日: 2016-09-07, 最終更新日: 2024-03-06)

主引用文献

Wu, Y.,Boxer, S.G.
A Critical Test of the Electrostatic Contribution to Catalysis with Noncanonical Amino Acids in Ketosteroid Isomerase.
J.Am.Chem.Soc., 138:11890-11895, 2016

PubMed Abstract: The vibrational Stark effect (VSE) has been used to measure the electric field in the active site of ketosteroid isomerase (KSI). These measured fields correlate with ΔG(⧧) in a series of conventional mutants, yielding an estimate for the electrostatic contribution to catalysis (Fried et al. Science 2014, 346, 1510-1513). In this work we test this result with much more conservative variants in which individual Tyr residues in the active site are replaced by 3-chlorotyrosine via amber suppression. The electric fields sensed at the position of the carbonyl bond involved in charge displacement during catalysis were characterized using the VSE, where the field sensitivity has been calibrated by vibrational Stark spectroscopy, solvatochromism, and MD simulations. A linear relationship is observed between the electric field and ΔG(⧧) that interpolates between wild-type and more drastic conventional mutations, reinforcing the evaluation of the electrostatic contribution to catalysis in KSI. A simplified model and calculation are developed to estimate changes in the electric field accompanying changes in the extended hydrogen-bond network in the active site. The results are consistent with a model in which the O-H group of a key active site tyrosine functions by imposing a static electrostatic potential onto the carbonyl bond. The model suggests that the contribution to catalysis from the active site hydrogen bonds is of similar weight to the distal interactions from the rest of the protein. A similar linear correlation was also observed between the proton affinity of KSI's active site and the catalytic rate, suggesting a direct connection between the strength of the H-bond and the electric field it exerts.

PubMed: 27545569
DOI: 10.1021/jacs.6b06843

実験手法

X-RAY DIFFRACTION (1.706 Å)