Loughborough University
Browse

Localising individual atoms of tryptophan side chains in the metallo-β-lactamase IMP-1 by pseudocontact shifts from paramagnetic lanthanoid tags at multiple sites

Download (4.84 MB)
journal contribution
posted on 2022-03-10, 14:50 authored by Henry W Orton, Iresha D Herath, Ansis Maleckis, Shereen Jabar, Monika Szabo, Bim Graham, Colum Breen, Lydia Topping, Stephen ButlerStephen Butler, Gottfried Otting
The metallo-β-lactamase IMP-1 features a flexible loop near the active site that assumes different conformations in single crystal structures, which may assist in substrate binding and enzymatic activity. To probe the position of this loop, we labelled the tryptophan residues of IMP-1 with 7-13C-indole and the protein with lanthanoid tags at three different sites. The magnetic susceptibility anisotropy (Δχ) tensors were determined by measuring pseudocontact shifts (PCSs) of backbone amide protons. The Δχ tensors were subsequently used to identify the atomic coordinates of the tryptophan side chains in the protein. The PCSs were sufficient to determine the location of Trp28, which is in the active site loop targeted by our experiments, with high accuracy. Its average atomic coordinates showed barely significant changes in response to the inhibitor captopril. It was found that localisation spaces could be defined with better accuracy by including only the PCSs of a single paramagnetic lanthanoid ion for each tag and tagging site. The effect was attributed to the shallow angle with which PCS isosurfaces tend to intersect if generated by tags and tagging sites that are identical except for the paramagnetic lanthanoid ion.

Funding

Australian Research Council (grant nos. CE200100012 and FL170100019)

European Regional Development Fund (grant no. 1.1.1.2/VIAA/2/18/381)

History

School

  • Science

Department

  • Chemistry

Published in

Magnetic Resonance

Volume

3

Issue

1

Pages

1 - 13

Publisher

Copernicus Publications

Version

  • VoR (Version of Record)

Rights holder

© The Authors

Publisher statement

This is an Open Access Article. It is published by Copernicus Publications under the Creative Commons Attribution 4.0 International Licence (CC BY 4.0). Full details of this licence are available at: https://creativecommons.org/licenses/by/4.0/

Acceptance date

2021-12-21

Publication date

2022-01-04

Copyright date

2022

eISSN

2699-0016

Language

  • en

Depositor

Dr Stephen Butler. Deposit date: 9 March 2022

Usage metrics

    Loughborough Publications

    Categories

    No categories selected

    Keywords

    Licence

    Exports

    RefWorks
    BibTeX
    Ref. manager
    Endnote
    DataCite
    NLM
    DC