Nuclear magnetic resonance-based determination of dioxygen binding sites in protein cavities

TY - JOUR

T1 - Nuclear magnetic resonance-based determination of dioxygen binding sites in protein cavities

AU - Kitahara, Ryo

AU - Sakuraba, Shun

AU - Kameda, Tomoshi

AU - Okuda, Sanshiro

AU - Xue, Mengjun

AU - Mulder, Frans A A

N1 - © 2017 The Protein Society.

PY - 2018/3/1

Y1 - 2018/3/1

N2 - The location and ligand accessibility of internal cavities in cysteine-free wild-type T4 lysozyme was investigated using O 2 gas-pressure NMR spectroscopy and molecular dynamics (MD) simulation. Upon increasing the concentration of dissolved O 2 in solvent to 8.9 mM, O 2-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 O 2 binding sites and their atomic coordinates from the 1/r 6 distance dependence of the PREs, we established an analytical procedure using Akaike's Information Criterion, in combination with a grid-search. Two O 2-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 O 2 revealed dioxygen ingress and egress as well as rotational and translational motions of O 2 in the cavities. It is therefore suggested that conformational fluctuations within the ground-state ensemble transiently develop channels for O 2 association with the internal protein cavities.

AB - The location and ligand accessibility of internal cavities in cysteine-free wild-type T4 lysozyme was investigated using O 2 gas-pressure NMR spectroscopy and molecular dynamics (MD) simulation. Upon increasing the concentration of dissolved O 2 in solvent to 8.9 mM, O 2-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 O 2 binding sites and their atomic coordinates from the 1/r 6 distance dependence of the PREs, we established an analytical procedure using Akaike's Information Criterion, in combination with a grid-search. Two O 2-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 O 2 revealed dioxygen ingress and egress as well as rotational and translational motions of O 2 in the cavities. It is therefore suggested that conformational fluctuations within the ground-state ensemble transiently develop channels for O 2 association with the internal protein cavities.

KW - Journal Article

UR - http://www.scopus.com/inward/record.url?scp=85042237395&partnerID=8YFLogxK

U2 - 10.1002/pro.3371

DO - 10.1002/pro.3371

M3 - Journal article

C2 - 29271012

SN - 0961-8368

VL - 27

SP - 769

EP - 779

JO - Protein Science

JF - Protein Science

IS - 3

ER -