NMR line shape analysis of a multi-state ligand binding mechanism in chitosanase

Chitosan interaction with chitosanase was examined through analysis of spectral line shapes in the NMR HSQC titration experiments. We established that the substrate, chitosan hexamer, binds to the enzyme through the three-state induced-fit mechanism with fast formation of the encounter complex follo...

Full description

Saved in:
Bibliographic Details
Published in:Journal of biomolecular NMR 2017-04, Vol.67 (4), p.309-319
Main Authors: Shinya, Shoko, Ghinet, Mariana G., Brzezinski, Ryszard, Furuita, Kyoko, Kojima, Chojiro, Shah, Sneha, Kovrigin, Evgenii L., Fukamizo, Tamo
Format: Article
Language:English
Subjects:
Bacterial Proteins - chemistry
Bacterial Proteins - metabolism
Binding Sites
Biochemistry
Biological and Medical Physics
Biophysics
Chitosan
Chitosan - chemistry
Chitosan - metabolism
Chitosanase
Conformation
Glycoside hydrolase
Glycoside Hydrolases - chemistry
Glycoside Hydrolases - metabolism
Isomerization
Kinetics
Ligands
Mapping
Models, Molecular
NMR
Nuclear magnetic resonance
Nuclear Magnetic Resonance, Biomolecular - methods
Physics
Physics and Astronomy
Protein Binding
Protein Structure, Tertiary
Spectroscopy/Spectrometry
Titration
Citations: Items that this one cites
Items that cite this one
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary:Chitosan interaction with chitosanase was examined through analysis of spectral line shapes in the NMR HSQC titration experiments. We established that the substrate, chitosan hexamer, binds to the enzyme through the three-state induced-fit mechanism with fast formation of the encounter complex followed by slow isomerization of the bound-state into the final conformation. Mapping of the chemical shift perturbations in two sequential steps of the mechanism highlighted involvement of the substrate-binding subsites and the hinge region in the binding reaction. Equilibrium parameters of the three-state model agreed with the overall thermodynamic dissociation constant determined by ITC. This study presented the first kinetic evidence of the induced-fit mechanism in the glycoside hydrolases.
ISSN:0925-2738
1573-5001
DOI:10.1007/s10858-017-0109-6