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Ligand-induced structural transitions combined with paramagnetic ions facilitate unambiguous NMR assignments of methyl groups in large proteins

NMR spectroscopy allows the study of biomolecules in close-to-native conditions. Structural information can be inferred from the NMR spectra when an assignment is available. Protein assignment is usually a time-consuming task, being specially challenging in the case of large, supramolecular systems....

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Autores principales: Mühlberg, Lars, Alarcin, Tuncay, Maass, Thorben, Creutznacher, Robert, Küchler, Richard, Mallagaray, Alvaro
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Springer Netherlands 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9247001/
https://www.ncbi.nlm.nih.gov/pubmed/35397749
http://dx.doi.org/10.1007/s10858-022-00394-0
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author Mühlberg, Lars
Alarcin, Tuncay
Maass, Thorben
Creutznacher, Robert
Küchler, Richard
Mallagaray, Alvaro
author_facet Mühlberg, Lars
Alarcin, Tuncay
Maass, Thorben
Creutznacher, Robert
Küchler, Richard
Mallagaray, Alvaro
author_sort Mühlberg, Lars
collection PubMed
description NMR spectroscopy allows the study of biomolecules in close-to-native conditions. Structural information can be inferred from the NMR spectra when an assignment is available. Protein assignment is usually a time-consuming task, being specially challenging in the case of large, supramolecular systems. Here, we present an extension of existing state-of-the-art strategies for methyl group assignment that partially overcomes signal overlapping and other difficulties associated to isolated methyl groups. Our approach exploits the ability of proteins to populate two or more conformational states, allowing for unique NOE restraints in each protein conformer. The method is compatible with automated assignment algorithms, granting assignments beyond the limits of a single protein state. The approach also benefits from long-range structural restraints obtained from metal-induced pseudocontact shifts (PCS) and paramagnetic relaxation enhancements (PREs). We illustrate the method with the complete assignment of the 199 methyl groups of a MIL(proS)V(proS)AT methyl-labeled sample of the UDP-glucose pyrophosphorylase enzyme from Leishmania major (LmUGP). Protozoan parasites of the genus Leishmania causes Leishmaniasis, a neglected disease affecting over 12 million people worldwide. LmUGP is responsible for the de novo biosynthesis of uridine diphosphate-glucose, a precursor in the biosynthesis of the dense surface glycocalyx involved in parasite survival and infectivity. NMR experiments with LmUGP and related enzymes have the potential to unravel new insights in the host resistance mechanisms used by Leishmania major. Our efforts will help in the development of selective and efficient drugs against Leishmania. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s10858-022-00394-0.
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spelling pubmed-92470012022-07-02 Ligand-induced structural transitions combined with paramagnetic ions facilitate unambiguous NMR assignments of methyl groups in large proteins Mühlberg, Lars Alarcin, Tuncay Maass, Thorben Creutznacher, Robert Küchler, Richard Mallagaray, Alvaro J Biomol NMR Article NMR spectroscopy allows the study of biomolecules in close-to-native conditions. Structural information can be inferred from the NMR spectra when an assignment is available. Protein assignment is usually a time-consuming task, being specially challenging in the case of large, supramolecular systems. Here, we present an extension of existing state-of-the-art strategies for methyl group assignment that partially overcomes signal overlapping and other difficulties associated to isolated methyl groups. Our approach exploits the ability of proteins to populate two or more conformational states, allowing for unique NOE restraints in each protein conformer. The method is compatible with automated assignment algorithms, granting assignments beyond the limits of a single protein state. The approach also benefits from long-range structural restraints obtained from metal-induced pseudocontact shifts (PCS) and paramagnetic relaxation enhancements (PREs). We illustrate the method with the complete assignment of the 199 methyl groups of a MIL(proS)V(proS)AT methyl-labeled sample of the UDP-glucose pyrophosphorylase enzyme from Leishmania major (LmUGP). Protozoan parasites of the genus Leishmania causes Leishmaniasis, a neglected disease affecting over 12 million people worldwide. LmUGP is responsible for the de novo biosynthesis of uridine diphosphate-glucose, a precursor in the biosynthesis of the dense surface glycocalyx involved in parasite survival and infectivity. NMR experiments with LmUGP and related enzymes have the potential to unravel new insights in the host resistance mechanisms used by Leishmania major. Our efforts will help in the development of selective and efficient drugs against Leishmania. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s10858-022-00394-0. Springer Netherlands 2022-04-10 2022 /pmc/articles/PMC9247001/ /pubmed/35397749 http://dx.doi.org/10.1007/s10858-022-00394-0 Text en © The Author(s) 2022 https://creativecommons.org/licenses/by/4.0/Open AccessThis article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/ (https://creativecommons.org/licenses/by/4.0/) .
spellingShingle Article
Mühlberg, Lars
Alarcin, Tuncay
Maass, Thorben
Creutznacher, Robert
Küchler, Richard
Mallagaray, Alvaro
Ligand-induced structural transitions combined with paramagnetic ions facilitate unambiguous NMR assignments of methyl groups in large proteins
title Ligand-induced structural transitions combined with paramagnetic ions facilitate unambiguous NMR assignments of methyl groups in large proteins
title_full Ligand-induced structural transitions combined with paramagnetic ions facilitate unambiguous NMR assignments of methyl groups in large proteins
title_fullStr Ligand-induced structural transitions combined with paramagnetic ions facilitate unambiguous NMR assignments of methyl groups in large proteins
title_full_unstemmed Ligand-induced structural transitions combined with paramagnetic ions facilitate unambiguous NMR assignments of methyl groups in large proteins
title_short Ligand-induced structural transitions combined with paramagnetic ions facilitate unambiguous NMR assignments of methyl groups in large proteins
title_sort ligand-induced structural transitions combined with paramagnetic ions facilitate unambiguous nmr assignments of methyl groups in large proteins
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9247001/
https://www.ncbi.nlm.nih.gov/pubmed/35397749
http://dx.doi.org/10.1007/s10858-022-00394-0
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