Related ArticlesNitroxide spin-labeled peptides for DNP-NMR in-cell studies.
FASEB J. 2019 10;33(10):11021-11027
Authors: Sani MA, Zhu S, Hofferek V, Separovic F
Abstract
Antimicrobial peptides (AMPs) that target lipid membranes show promise as alternatives to conventional antibiotics. However, the molecular mechanisms of membrane perturbation, as most studies are performed in model systems and in-cell structural studies, have yet to be achieved. Solid-state NMR spectroscopy is a valuable technique to investigate peptide-membrane interactions and to determine the structure of peptides, but the short lifespan of bacteria, especially under magic angle spinning conditions, has not permitted in-cell structural studies. Here, we present the first dynamic nuclear polarization (DNP)-NMR in-cell studies of Escherichia coli bacteria incubated with the AMP maculatin 1.1 (Mac1) in combination with novel nitroxide spin-labeled peptides 2,2,6,6-tetramethylpiperidine-N-oxyl-4-amino-4-carboxylic acid (TOAC)-[F3W]-Mac1 (MacW) and TOAC-TOAC-MacW. The in-cell 13C and 15N signal NMR enhancements, and 1H spin-lattice T1 relaxation times showed that TOAC-MacW and TOAC-TOAC-MacW performed better than the more hydrophilic biradical AMUPol used for DNP studies. Furthermore, the pores formed by the AMP increased the signal enhancements and decreased T1 values of specifically 13C- and 15N-labeled Mac1. This approach has a great potential for determining the first in situ structures of AMPs in bacteria.-Sani, M.-A., Zhu, S., Hofferek, V., Separovic, F. Nitroxide spin-labeled peptides for DNP-NMR in-cell studies.
[NMR paper] A bioresistant nitroxide spin label for in-cell EPR spectroscopy: in vitro and in oocytes protein structural dynamics studies.
A bioresistant nitroxide spin label for in-cell EPR spectroscopy: in vitro and in oocytes protein structural dynamics studies.
Approaching proteins structural dynamics and protein-protein interactions in the cellular environment is a fundamental challenge. Due to its absolute sensitivity and to its selectivity to paramagnetic species, Site-Directed Spin Labeling (SDSL) combined with Electron Paramagnetic Resonance (EPR) has the potential to evolve into an efficient method to follow conformational changes in proteins directly inside cells. Until now, the use of nitroxyde-based spin labels...
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12-12-2017 02:12 AM
High-field ELDOR-detected NMR study of a nitroxide radical in disordered solids: Towards characterization of heterogeneity of microenvironments in spin-labeled systems
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High-field ELDOR-detected NMR study of a nitroxide radical in disordered solids: Towards characterization of heterogeneity of microenvironments in spin-labeled systems
With nitroxides one of the most popular polarizing agents for DNP-NMR spectroscopy, this article gives more insights into the precise interactions (hyperfine, quadrupolar, g anisotropy ...) at high magnetic fields.
Nalepa, A., et al., High-field ELDOR-detected NMR study of a nitroxide radical in disordered solids: Towards characterization of heterogeneity of microenvironments in...
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08-04-2014 11:16 PM
[NMR paper] High-field ELDOR-detected NMR study of a nitroxide radical in disordered solids: Towards characterization of heterogeneity of microenvironments in spin-labeled systems.
High-field ELDOR-detected NMR study of a nitroxide radical in disordered solids: Towards characterization of heterogeneity of microenvironments in spin-labeled systems.
High-field ELDOR-detected NMR study of a nitroxide radical in disordered solids: Towards characterization of heterogeneity of microenvironments in spin-labeled systems.
J Magn Reson. 2014 Mar 12;242C:203-213
Authors: Nalepa A, Möbius K, Lubitz W, Savitsky A
Abstract
The combination of high-field EPR with site-directed spin-labeling (SDSL) techniques employing...
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04-02-2014 11:54 PM
High-field ELDOR-detected NMR study of a nitroxide radical in disordered solids:Towards characterization of heterogeneity of microenvironments in spin-labeled systems
High-field ELDOR-detected NMR study of a nitroxide radical in disordered solids:Towards characterization of heterogeneity of microenvironments in spin-labeled systems
Publication date: Available online 12 March 2014
Source:Journal of Magnetic Resonance</br>
Author(s): Anna Nalepa , Klaus Möbius , Wolfgang Lubitz , Anton Savitsky</br>
The combination of high-field EPR with site-directed spin-labeling (SDSL) techniques employing nitroxide radicals has turned out to be particularly powerful in probing the polarity and proticity characteristics of protein/matrix...
[NMR paper] A rapid procedure to isolate isotopically labeled peptides for NMR studies: application to the Disabled-2 sulfatide-binding motif.
A rapid procedure to isolate isotopically labeled peptides for NMR studies: application to the Disabled-2 sulfatide-binding motif.
Related Articles A rapid procedure to isolate isotopically labeled peptides for NMR studies: application to the Disabled-2 sulfatide-binding motif.
J Pept Sci. 2014 Jan 27;
Authors: Xiao S, Zhao X, Finkielstein CV, Capelluto DG
Abstract
A procedure for obtaining isotopically labeled peptides, by combining affinity chromatography, urea-equilibrated gel filtration, and hydrophobic chromatography procedures, is...
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01-29-2014 02:01 PM
[NMR paper] Cell-free synthesis of 15N-labeled proteins for NMR studies.
Cell-free synthesis of 15N-labeled proteins for NMR studies.
Related Articles Cell-free synthesis of 15N-labeled proteins for NMR studies.
IUBMB Life. 2005 Sep;57(9):615-22
Authors: Ozawa K, Dixon NE, Otting G
Modern cell-free in vitro protein synthesis systems present powerful tools for the synthesis of isotope-labeled proteins in high yields. The production of selectively 15 N-labeled proteins from 15 N-labeled amino acids is particularly economic and yields are often sufficient to analyze the proteins very quickly by two-dimensional NMR...
Nitroxide spin-labeled peptides for DNP-NMR in-cell studies.
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