Articles | Volume 3, issue 1
https://doi.org/10.5194/mr-3-65-2022
© Author(s) 2022. This work is distributed under
the Creative Commons Attribution 4.0 License.
the Creative Commons Attribution 4.0 License.
https://doi.org/10.5194/mr-3-65-2022
© Author(s) 2022. This work is distributed under
the Creative Commons Attribution 4.0 License.
the Creative Commons Attribution 4.0 License.
Research article
| 
09 May 2022
Research article |
| 09 May 2022
| 09 May 2022
Localising nuclear spins by pseudocontact shifts from a single tagging site
Henry W. Orton
ARC Centre of Excellence for Innovations in Peptide & Protein Science, Research School of Chemistry, Australian National University, Canberra, ACT 2601, Australia
Elwy H. Abdelkader
ARC Centre of Excellence for Innovations in Peptide & Protein Science, Research School of Chemistry, Australian National University, Canberra, ACT 2601, Australia
Lydia Topping
Department of Chemistry, Loughborough University, Epinal Way, Loughborough, LE11 3TU, United Kingdom
Stephen J. Butler
Department of Chemistry, Loughborough University, Epinal Way, Loughborough, LE11 3TU, United Kingdom
Gottfried Otting
CORRESPONDING AUTHOR
ARC Centre of Excellence for Innovations in Peptide & Protein Science, Research School of Chemistry, Australian National University, Canberra, ACT 2601, Australia
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Henry W. Orton, Iresha D. Herath, Ansis Maleckis, Shereen Jabar, Monika Szabo, Bim Graham, Colum Breen, Lydia Topping, Stephen J. Butler, and Gottfried Otting
Magn. Reson., 3, 1–13, https://doi.org/10.5194/mr-3-1-2022, https://doi.org/10.5194/mr-3-1-2022, 2022
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This paper explores a method for determining the solution structure of a solvent-exposed polypeptide segment (the L3 loop), which is next to the active site of the penicillin-degrading enzyme IMP-1. Tagging three different sites on the protein with paramagnetic metal ions allowed positioning of the L3 loop with atomic resolution. It was found that the method was more robust when omitting data obtained with different metal ions if obtained with the same tag at the same tagging site.
Henry William Orton, Thomas Huber, and Gottfried Otting
Magn. Reson., 1, 1–12, https://doi.org/10.5194/mr-1-1-2020, https://doi.org/10.5194/mr-1-1-2020, 2020
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Nuclear magnetic resonance is a technique that allows the measurement of the nanoscale distances between the atoms of a molecule. It is often relied upon for finding the structure of a molecule and how atoms are bonded, but measurements become inaccurate for large distances and therefore for large biological molecules. This research presents a new software for calculating the distances between nuclei and unpaired electrons which offers more accurate long-range distances in biological molecules.
Damian Van Raad, Gottfried Otting, and Thomas Huber
Magn. Reson., 4, 187–197, https://doi.org/10.5194/mr-4-187-2023, https://doi.org/10.5194/mr-4-187-2023, 2023
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A novel cell-free protein synthesis system called eCells produces amino acids based on specific isotopes using low-cost precursors. The system selectively labels methyl groups, i.e valine and leucine, with high efficiency. eCells achieve high levels of 13C incorporation and deuteration in protein preparations, making them suitable for NMR experiments of large protein complexes. They are easy to prepare, can be scaled up in volume and are a promising tool for protein production and NMR studies.
Sreelakshmi Mekkattu Tharayil, Mithun C. Mahawaththa, Akiva Feintuch, Ansis Maleckis, Sven Ullrich, Richard Morewood, Michael J. Maxwell, Thomas Huber, Christoph Nitsche, Daniella Goldfarb, and Gottfried Otting
Magn. Reson., 3, 169–182, https://doi.org/10.5194/mr-3-169-2022, https://doi.org/10.5194/mr-3-169-2022, 2022
Short summary
Short summary
Having shown that tagging a protein at a single site with different lanthanoid complexes delivers outstanding structural information at a selected site of a protein (such as active sites and ligand binding sites), we now present a simple way by which different lanthanoid complexes can be assembled on a highly solvent-exposed cysteine residue. Furthermore, the chemical assembly is selective for selenocysteine, if a selenocysteine residue can be introduced into the protein of interest.
Henry W. Orton, Iresha D. Herath, Ansis Maleckis, Shereen Jabar, Monika Szabo, Bim Graham, Colum Breen, Lydia Topping, Stephen J. Butler, and Gottfried Otting
Magn. Reson., 3, 1–13, https://doi.org/10.5194/mr-3-1-2022, https://doi.org/10.5194/mr-3-1-2022, 2022
Short summary
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This paper explores a method for determining the solution structure of a solvent-exposed polypeptide segment (the L3 loop), which is next to the active site of the penicillin-degrading enzyme IMP-1. Tagging three different sites on the protein with paramagnetic metal ions allowed positioning of the L3 loop with atomic resolution. It was found that the method was more robust when omitting data obtained with different metal ions if obtained with the same tag at the same tagging site.
Sreelakshmi Mekkattu Tharayil, Mithun Chamikara Mahawaththa, Choy-Theng Loh, Ibidolapo Adekoya, and Gottfried Otting
Magn. Reson., 2, 1–13, https://doi.org/10.5194/mr-2-1-2021, https://doi.org/10.5194/mr-2-1-2021, 2021
Short summary
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A new way is presented for creating lanthanide binding sites on proteins using site-specifically introduced phosphoserine residues. The paramagnetic effects of lanthanides generate long-range effects, which contain structural information and are readily measured by NMR spectroscopy. Excellent correlations between experimentally observed and back-calculated pseudocontact shifts attest to very good immobilization of the lanthanide ions relative to the proteins.
Henry William Orton, Thomas Huber, and Gottfried Otting
Magn. Reson., 1, 1–12, https://doi.org/10.5194/mr-1-1-2020, https://doi.org/10.5194/mr-1-1-2020, 2020
Short summary
Short summary
Nuclear magnetic resonance is a technique that allows the measurement of the nanoscale distances between the atoms of a molecule. It is often relied upon for finding the structure of a molecule and how atoms are bonded, but measurements become inaccurate for large distances and therefore for large biological molecules. This research presents a new software for calculating the distances between nuclei and unpaired electrons which offers more accurate long-range distances in biological molecules.
Related subject area
Field: Liquid-state NMR | Topic: Applications – biological macromolecules
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The 20 genetically encoded amino acids provide the basis for most proteins and peptides that make up the machinery of life. This limited repertoire is vastly expanded by the introduction of non-canonical amino acids (ncAAs). Studying the structure of protein-containing ncAAs requires new computational representations that are compatible with existing modelling software. We have developed an online tool for this to aid future structural studies of this class of complex biopolymer.
Sreelakshmi Mekkattu Tharayil, Mithun C. Mahawaththa, Akiva Feintuch, Ansis Maleckis, Sven Ullrich, Richard Morewood, Michael J. Maxwell, Thomas Huber, Christoph Nitsche, Daniella Goldfarb, and Gottfried Otting
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Stephan Grzesiek, Johannes Paladini, Judith Habazettl, and Rajesh Sonti
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We show here that binding of the anticancer drug imatinib to the ATP site of Abelson kinase and not binding to its allosteric site coincides with the opening of the kinase regulatory core at nanomolar concentrations. This has implications for the understanding of Abelson’s kinase regulation and activity during medication as well as for the design of new Abelson kinase inhibitors.
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The overall aim of the study is to provide a basis from which to improve the ability of tamoxifen family drugs to reduce the activity of a secondary target protein, calmodulin, during tumour development. The main conclusion is that the binding of a tamoxifen analogue is quite unlike that of other anti-calmodulin compounds in that two drug molecules bring the two domains of calmodulin into close proximity, but they are not fixed in orientation relative to the protein.
Chih-Ting Huang, Yei-Chen Lai, Szu-Yun Chen, Meng-Ru Ho, Yun-Wei Chiang, and Shang-Te Danny Hsu
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The iron–sulfur cluster binding properties of human mitoNEET have been investigated by 1D and 2D 1H paramagnetic NMR spectroscopy. The NMR spectra of both oxidized and reduced mitoNEET are significantly different from those reported previously for other [Fe2S2] proteins. Our findings revealed the unique electronic properties of mitoNEET and suggests that the specific electronic structure of the cluster possibly drives the functional properties of different [Fe2S2] proteins.
Mitsuhiro Takeda, Yohei Miyanoiri, Tsutomu Terauchi, and Masatsune Kainosho
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Although both the hydrophobic aliphatic chain and hydrophilic ζ-amino group of the lysine side chain presumably contribute to the structures and functions of proteins, the dual nature of the lysine residue has not been fully understood yet, due to the lack of appropriate methods to acquire comprehensive information on its long consecutive methylene chain at the atomic scale. We describe herein a novel strategy to address the current situation using nuclear magnetic resonance spectroscopy.
Xingjian Xu, Igor Dikiy, Matthew R. Evans, Leandro P. Marcelino, and Kevin H. Gardner
Magn. Reson., 2, 63–76, https://doi.org/10.5194/mr-2-63-2021, https://doi.org/10.5194/mr-2-63-2021, 2021
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While most proteins adopt one conformation, several interconvert between two or more very different structures. Knowing how sequence changes and small-molecule binding can control this behavior is essential for both understanding biology and inspiring new “molecular switches” which can control cellular pathways. This work contributes by examining these topics in the ARNT protein, showing that features of both the folded and unfolded states contribute to the interconversion process.
Rubin Dasgupta, Karthick B. S. S. Gupta, Huub J. M. de Groot, and Marcellus Ubbink
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A method is demonstrated that can help in sequence-specific NMR signal assignment to nuclear spins near a strongly paramagnetic metal in an enzyme. A combination of paramagnetically tailored NMR experiments and second-shell mutagenesis was used to attribute previously observed chemical exchange processes in the active site of laccase to specific histidine ligands. The signals of nuclei close to the metal can be used as spies to unravel the role of motions in the catalytic process.
Sreelakshmi Mekkattu Tharayil, Mithun Chamikara Mahawaththa, Choy-Theng Loh, Ibidolapo Adekoya, and Gottfried Otting
Magn. Reson., 2, 1–13, https://doi.org/10.5194/mr-2-1-2021, https://doi.org/10.5194/mr-2-1-2021, 2021
Short summary
Short summary
A new way is presented for creating lanthanide binding sites on proteins using site-specifically introduced phosphoserine residues. The paramagnetic effects of lanthanides generate long-range effects, which contain structural information and are readily measured by NMR spectroscopy. Excellent correlations between experimentally observed and back-calculated pseudocontact shifts attest to very good immobilization of the lanthanide ions relative to the proteins.
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Short summary
Installing a tag containing a paramagnetic metal ion on a protein can lead to large changes (pseudocontact shifts) in the resonances observed in NMR spectra. These are easily measured and contain valuable long-range structural information. The present work shows that a single tagging site furnished with different tags can be sufficient to localise atoms in proteins with high accuracy. In fact, this strategy works almost as well as the same number of tags distributed over multiple tagging sites.
Installing a tag containing a paramagnetic metal ion on a protein can lead to large changes...
