| Zdroj: |
Kitahara, R, Sakuraba, S, Kameda, T, Okuda, S, Xue, M & Mulder, F A A 2018, ' Nuclear magnetic resonance-based determination of dioxygen binding sites in protein cavities ', Protein Science, vol. 27, no. 3, pp. 769-779 . https://doi.org/10.1002/pro.3371 |
| Popis: |
The location and ligand accessibility of internal cavities in cysteine-free wild-type T4 lysozyme was investigated using O2 gas-pressure NMR spectroscopy and molecular dynamics (MD) simulation. Upon increasing the concentration of dissolved O2 in solvent to 8.9 mM, O2 -induced paramagnetic relaxation enhancements (PREs) to the backbone amide and side chain methyl protons were observed, specifically around two cavities in the C-terminal domain. To determine the number of O2 binding sites and their atomic coordinates from the 1/r6 distance dependence of the PREs, we established an analytical procedure using Akaike's Information Criterion, in combination with a grid-search. Two O2 -accessible sites were identified in internal cavities: One site was consistent with the xenon-binding site in the protein in crystal, and the other site was established to be a novel ligand-binding site. MD simulations performed at 10 and 100 mM O2 revealed dioxygen ingress and egress as well as rotational and translational motions of O2 in the cavities. It is therefore suggested that conformational fluctuations within the ground-state ensemble transiently develop channels for O2 association with the internal protein cavities. This article is protected by copyright. All rights reserved. |
