How to tackle protein structural data from solution and solid state: An integrated approach

Carlon, Azzurra; Ravera, Enrico; Andrałojć, Witold; Parigi, Giacomo; Murshudov, Garib N.; Luchinat, Claudio

doi:10.1016/j.pnmrs.2016.01.001

Cited by 28 publications

(26 citation statements)

References 196 publications

(199 reference statements)

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“…It is well established that chemical shift frequencies of individual resonances are affected by the local electronic environment, and can thus provide local structural and dynamic information. [31][32][33][34] In particular, the binding of metal ions to specific residues affects the NMR resonances of proximate residues. Since Cu 2 + has unpaired electrons, the associated paramagnetic effects strongly influence both the chemical shifts and the relaxation rates of resonances from nearby sites.…”

Section: Resultsmentioning

confidence: 99%

Metal‐ion Binding to Host Defense Peptide Piscidin 3 Observed in Phospholipid Bilayers by Magic Angle Spinning Solid‐state NMR

et al. 2018

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Cationic antimicrobial peptides (AMPs) are essential components of the innate immune system. They have attracted interest as novel compounds with the potential to treat infections associated with multi‐drug resistant bacteria. In this study, we investigate piscidin 3 (P3), an AMP that was first discovered in the mast cells of hybrid striped bass. Prior studies showed that P3 is less active than its homolog piscidin 1 (P1) against planktonic bacteria. However, P3 has the advantage of being less toxic to mammalian cells and more active on biofilms and persister cells. Both P1 and P3 cross bacterial membranes and co‐localize with intracellular DNA but P3 is more condensing to DNA while P1 is more membrane active. Recently, we showed that both peptides coordinate Cu2+ through an amino‐terminal copper and nickel (ATCUN) motif. We also demonstrated that the bactericidal effects of P3 are linked to its ability to form radicals that nick DNA in the presence of Cu2+. Since metal binding and membrane crossing by P3 is biologically important, we apply in this study solid‐state NMR spectroscopy to uniformly 13C‐15N‐labeled peptide samples to structurally characterize the ATCUN motif of P3 bound to bilayers and coordinated to Ni2+ and Cu2+. These experiments are supplemented with density functional theory calculations. Taken together, these studies refine the arrangement of not only the backbone but also side chain atoms of an AMP simultaneously bound to metal ions and phospholipid bilayers.

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Section: Resultsmentioning

confidence: 99%

Metal‐ion Binding to Host Defense Peptide Piscidin 3 Observed in Phospholipid Bilayers by Magic Angle Spinning Solid‐state NMR

et al. 2018

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“…Owing to their ‘global’ character, PCSs and RDCs represent the best candidates for shedding light on the longstanding issue of the combination of structural knowledge derived from solution and solid-state (crystal) data (Carlon, Ravera, Andrałojć et al , 2016). NMR data, indeed, are often used in comparison (or, ideally, in combination) with crystal diffraction data, whenever available.…”

Section: Nmr Restraints In Refinementmentioning

confidence: 99%

“…Let us consider the refinement of a ternary Sxl–Unr–msl2-mRNA regulatory complex (Carlon, Ravera, Hennig et al , 2016). This complex consists of both RNA-recognition motifs (RRMs) of Sxl, the first of five cold-shock domains of Unr (CSD1) and an 18-mer single-stranded RNA derived from msl2-mRNA.…”

Section: Nmr Restraints In Refinementmentioning

confidence: 99%

Overview of refinement procedures withinREFMAC5: utilizing data from different sources

Kovalevskiy

Nicholls

Long

et al. 2018

Acta Crystallogr D Struct Biol

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224

194

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Here, a macromolecule-centred approach to three-dimensional structure determination as implemented in REFMAC5 is considered. The use of restraints to transfer chemical and structural information during macromolecular refinement, and how different sources of information can be combined in order to achieve models that are more consistent with data derived from a variety of experimental techniques, including macromolecular crystallography, cryo-EM and NMR spectroscopy, are discussed.

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“…It should be noted that that the structural characterization of ensembles of conformations is an ill-posed inverse problem since the NMR observables are ensemble averages. Thus, there is an infinite number of ensembles of structures that can satisfy the experimental data equally well [9]. Additional modeling is required to better characterize the conformational space.…”

Section: Introductionmentioning

confidence: 99%

The MPS1 kinase NTE region has helical propensity and preferred conformations towards the TPR domain

Hiruma

Matsoukas

Touw

et al. 2020

Preprint

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The mitotic spindle assembly checkpoint (SAC) ensures accurate segregation of chromosomes by preventing onset of anaphase until all chromosomes are properly attached to spindle microtubules. The Monopolar spindle 1 (MPS1) kinase is one of the SAC components, localizing at unattached kinetochores by an N-terminal localization module. This module comprises a flexible NTE module and the TPR domain, which we previously characterized for their contribution to kinetochore binding. Here we discuss the conformations of the highly flexible NTE with respect to the TPR domain, using paramagnetic NMR. The distance restraints derived from paramagnetic relaxation enhancements (PREs) show that the mobile NTE can be found in proximity of a large but specific part of the surface area of the TPR domain. To sample the conformational space of the NTE in the context of the NTE-TPR module, we used the ab initio Rosetta approach supplemented by paramagnetic NMR restraints. We find that many NTE residues have a propensity to form helical structures and that the module localizes at the convex surface of the TPR domain. This work demonstrates the highly dynamic nature of the interactions between the NTE and TPR domains and it shows that the convex rather than the canonical concave TPR surface mediates interactions, leading to the auto-inhibition that the TPR exerts upon the NTE region in the context of SAC signaling. 144Total 7 132 170 309

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How to tackle protein structural data from solution and solid state: An integrated approach

Cited by 28 publications

References 196 publications

Metal‐ion Binding to Host Defense Peptide Piscidin 3 Observed in Phospholipid Bilayers by Magic Angle Spinning Solid‐state NMR

Metal‐ion Binding to Host Defense Peptide Piscidin 3 Observed in Phospholipid Bilayers by Magic Angle Spinning Solid‐state NMR

Overview of refinement procedures withinREFMAC5: utilizing data from different sources

The MPS1 kinase NTE region has helical propensity and preferred conformations towards the TPR domain

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