scholarly journals Acquisition of ionic copper by a bacterial outer membrane protein

2020 ◽  
Author(s):  
Satya Prathyusha Bhamidimarri ◽  
Tessa R. Young ◽  
Muralidharan Shanmugam ◽  
Sandra Soderholm ◽  
Bastien Belzunces ◽  
...  
Keyword(s):  
Metal Binding ◽  
Paramagnetic Resonance ◽  
Copper Crystal ◽  
Ionic Copper ◽  
Icp Ms ◽  
Wide Range ◽  
Copper Storage ◽  
And Function ◽  
Electron Paramagnetic ◽  
Copper Import

AbstractCopper, while toxic in excess, is an essential micronutrient in all kingdoms of life due to its essential role in the structure and function of many proteins. Proteins mediating ionic copper import are known in eukaryotes, but have not yet been described in prokaryotes. Here we show that Pseudomonas aeruginosa OprC is a TonB-dependent transporter that mediates acquisition of ionic copper. Crystal structures of wild type and mutant OprC variants with silver and copper, as well as ICP-MS and electron paramagnetic resonance (EPR), suggest that binding of Cu(I) occurs via a surface-exposed methionine track leading towards an unprecedented CxxxM-HxM metal binding site that binds Cu(I) directly and can facilitate reduction of Cu(II) via the cysteine thiol. Together with quantitative proteomics and growth assays, our data identify OprC as an abundant component of bacterial copper biology that enables copper acquisition and potentially copper storage under a wide range of environmental conditions.

scholarly journals Acquisition of ionic copper by the bacterial outer membrane protein OprC through a novel binding site

PLoS Biology ◽  
2021 ◽  
Vol 19 (11) ◽  
pp. e3001446
Author(s):  
Satya Prathyusha Bhamidimarri ◽  
Tessa R. Young ◽  
Muralidharan Shanmugam ◽  
Sandra Soderholm ◽  
Arnaud Baslé ◽  
...  
Keyword(s):  
Binding Site ◽  
Outer Membrane ◽  
Metal Binding ◽  
Infection Model ◽  
Metal Binding Site ◽  
Ionic Copper ◽  
Wide Range ◽  
Bacterial Outer Membrane ◽  
And Function ◽  
Copper Import

Copper, while toxic in excess, is an essential micronutrient in all kingdoms of life due to its essential role in the structure and function of many proteins. Proteins mediating ionic copper import have been characterised in detail for eukaryotes, but much less so for prokaryotes. In particular, it is still unclear whether and how gram-negative bacteria acquire ionic copper. Here, we show that Pseudomonas aeruginosa OprC is an outer membrane, TonB-dependent transporter that is conserved in many Proteobacteria and which mediates acquisition of both reduced and oxidised ionic copper via an unprecedented CxxxM-HxM metal binding site. Crystal structures of wild-type and mutant OprC variants with silver and copper suggest that acquisition of Cu(I) occurs via a surface-exposed “methionine track” leading towards the principal metal binding site. Together with whole-cell copper quantitation and quantitative proteomics in a murine lung infection model, our data identify OprC as an abundant component of bacterial copper biology that may enable copper acquisition under a wide range of conditions.

Nanoscale lipid membrane mimetics in spin-labeling and electron paramagnetic resonance spectroscopy studies of protein structure and function

2017 ◽  
Vol 6 (1) ◽  
pp. 75-92 ◽  
Author(s):  
Elka R. Georgieva
Keyword(s):  
Electron Paramagnetic Resonance ◽  
Protein Structure ◽  
Membrane Proteins ◽  
Membrane Protein ◽  
Spin Labeling ◽  
Structure And Function ◽  
Protein Structure And Function ◽  
Paramagnetic Resonance ◽  
And Function ◽  
Electron Paramagnetic

AbstractCellular membranes and associated proteins play critical physiological roles in organisms from all life kingdoms. In many cases, malfunction of biological membranes triggered by changes in the lipid bilayer properties or membrane protein functional abnormalities lead to severe diseases. To understand in detail the processes that govern the life of cells and to control diseases, one of the major tasks in biological sciences is to learn how the membrane proteins function. To do so, a variety of biochemical and biophysical approaches have been used in molecular studies of membrane protein structure and function on the nanoscale. This review focuses on electron paramagnetic resonance with site-directed nitroxide spin-labeling (SDSL EPR), which is a rapidly expanding and powerful technique reporting on the local protein/spin-label dynamics and on large functionally important structural rearrangements. On the other hand, adequate to nanoscale study membrane mimetics have been developed and used in conjunction with SDSL EPR. Primarily, these mimetics include various liposomes, bicelles, and nanodiscs. This review provides a basic description of the EPR methods, continuous-wave and pulse, applied to spin-labeled proteins, and highlights several representative applications of EPR to liposome-, bicelle-, or nanodisc-reconstituted membrane proteins.

Biodistribution and pharmacokinetic studies of SPION using particle electron paramagnetic resonance, MRI and ICP-MS

Nanomedicine ◽  
2015 ◽  
Vol 10 (11) ◽  
pp. 1751-1760 ◽  
Author(s):  
Oliviero L Gobbo ◽  
Friedrich Wetterling ◽  
Peter Vaes ◽  
Stephanie Teughels ◽  
Farouk Markos ◽  
...  
Keyword(s):  
Electron Paramagnetic Resonance ◽  
Pharmacokinetic Studies ◽  
Paramagnetic Resonance ◽  
Icp Ms ◽  
Electron Paramagnetic

scholarly journals DeerLab: a comprehensive software package for analyzing dipolar electron paramagnetic resonance spectroscopy data

2020 ◽  
Vol 1 (2) ◽  
pp. 209-224 ◽  
Author(s):  
Luis Fábregas Ibáñez ◽  
Gunnar Jeschke ◽  
Stefan Stoll
Keyword(s):  
Electron Paramagnetic Resonance ◽  
Software Package ◽  
Method Development ◽  
Electron Paramagnetic Resonance Spectroscopy ◽  
Global Analysis ◽  
Resonance Spectroscopy ◽  
Paramagnetic Resonance ◽  
Least Squares Fit ◽  
Wide Range ◽  
Electron Paramagnetic

Abstract. Dipolar electron paramagnetic resonance (EPR) spectroscopy (DEER and other techniques) enables the structural characterization of macromolecular and biological systems by measurement of distance distributions between unpaired electrons on a nanometer scale. The inference of these distributions from the measured signals is challenging due to the ill-posed nature of the inverse problem. Existing analysis tools are scattered over several applications with specialized graphical user interfaces. This renders comparison, reproducibility, and method development difficult. To remedy this situation, we present DeerLab, an open-source software package for analyzing dipolar EPR data that is modular and implements a wide range of methods. We show that DeerLab can perform one-step analysis based on separable non-linear least squares, fit dipolar multi-pathway models to multi-pulse DEER data, run global analysis with non-parametric distributions, and use a bootstrapping approach to fully quantify the uncertainty in the analysis.

Ultra-broadband EPR spectroscopy in field and frequency domains

2018 ◽  
Vol 20 (22) ◽  
pp. 15528-15534 ◽  
Author(s):  
P. Neugebauer ◽  
D. Bloos ◽  
R. Marx ◽  
P. Lutz ◽  
M. Kern ◽  
...  
Keyword(s):  
Electron Paramagnetic Resonance ◽  
Magnetic Properties ◽  
Epr Spectroscopy ◽  
Paramagnetic Resonance ◽  
Powerful Technique ◽  
Electronic And Magnetic Properties ◽  
Wide Range ◽  
Frequency Domains ◽  
Electron Paramagnetic

Electron paramagnetic resonance (EPR) is a powerful technique to investigate the electronic and magnetic properties of a wide range of materials.

scholarly journals An EPR Study on the Interaction between the Cu(I) Metal Binding Domains of ATP7B and the Atox1 Metallochaperone

2020 ◽  
Vol 21 (15) ◽  
pp. 5536
Author(s):  
Michael Zaccak ◽  
Zena Qasem ◽  
Lada Gevorkyan-Airapetov ◽  
Sharon Ruthstein
Keyword(s):  
Metal Binding ◽  
Continuous Wave ◽  
Regulation System ◽  
Membrane Domain ◽  
Paramagnetic Resonance ◽  
Copper Trafficking ◽  
Binding Domains ◽  
Metal Binding Domain ◽  
Pulsed Electron Paramagnetic Resonance ◽  
Electron Paramagnetic

Copper’s essentiality and toxicity mean it requires a sophisticated regulation system for its acquisition, cellular distribution and excretion, which until now has remained elusive. Herein, we applied continuous wave (CW) and pulsed electron paramagnetic resonance (EPR) spectroscopy in solution to resolve the copper trafficking mechanism in humans, by considering the route travelled by Cu(I) from the metallochaperone Atox1 to the metal binding domains of ATP7B. Our study revealed that Cu(I) is most likely mediated by the binding of the Atox1 monomer to metal binding domain 1 (MBD1) and MBD4 of ATP7B in the final part of its extraction pathway, while the other MBDs mediate this interaction and participate in copper transfer between the various MBDs to the ATP7B membrane domain. This research also proposes that MBD1-3 and MBD4-6 act as two independent units.

scholarly journals In situ kinetic measurements of α-synuclein aggregation reveal large population of short-lived oligomers

PLoS ONE ◽  
2021 ◽  
Vol 16 (1) ◽  
pp. e0245548
Author(s):  
Enrico Zurlo ◽  
Pravin Kumar ◽  
Georg Meisl ◽  
Alexander J. Dear ◽  
Dipro Mondal ◽  
...  
Keyword(s):  
Self Assembly ◽  
Large Population ◽  
Paramagnetic Resonance ◽  
Kinetic Measurements ◽  
Toxic Species ◽  
Self Assembling ◽  
Wide Range ◽  
Electron Paramagnetic

Knowledge of the mechanisms of assembly of amyloid proteins into aggregates is of central importance in building an understanding of neurodegenerative disease. Given that oligomeric intermediates formed during the aggregation reaction are believed to be the major toxic species, methods to track such intermediates are clearly needed. Here we present a method, electron paramagnetic resonance (EPR), by which the amount of intermediates can be measured over the course of the aggregation, directly in the reacting solution, without the need for separation. We use this approach to investigate the aggregation of α-synuclein (αS), a synaptic protein implicated in Parkinson’s disease and find a large population of oligomeric species. Our results show that these are primary oligomers, formed directly from monomeric species, rather than oligomers formed by secondary nucleation processes, and that they are short-lived, the majority of them dissociates rather than converts to fibrils. As demonstrated here, EPR offers the means to detect such short-lived intermediate species directly in situ. As it relies only on the change in size of the detected species, it will be applicable to a wide range of self-assembling systems, making accessible the kinetics of intermediates and thus allowing the determination of their rates of formation and conversion, key processes in the self-assembly reaction.

Electron paramagnetic resonance study of the reactions of the spin trap 3,5-dibromo-4-nitrosobenzenesulfonate

1988 ◽  
Vol 66 (5) ◽  
pp. 1153-1158 ◽  
Author(s):  
Peter Smith ◽  
Jill Suzanne Robertson
Keyword(s):  
Electron Paramagnetic Resonance ◽  
Aqueous Solutions ◽  
Electron Paramagnetic Resonance Study ◽  
Oxidation Mechanism ◽  
Azo Compounds ◽  
Electronic Effects ◽  
Paramagnetic Resonance ◽  
Wide Range ◽  
Methacrylic Monomers ◽  
Electron Paramagnetic

Using rapid-mixing, continuous-flow TiCl3-based techniques and also by means of static-sample studies involving the thermal decomposition of symmetric aliphatic azo compounds, we have characterized by electron paramagnetic resonance 22 spin adducts of 3,5-dibromo-4-nitrosobenzenesulphonate, 1, in aqueous solution at 25 °C. These spin adducts, all nitroxides, exhibit a moderately wide range of a-nitrogen and β-CH proton splitting constants, which we discuss in terms of steric and electronic effects. In connection with these studies, blank experiments showed that aqueous solutions of 1 gave no radicals when exposed to light and heat. In addition, we have studied by electron paramagnetic resonance at 25 °C static samples of aqueous solutions of 1 both by itself and in the presence of each of several acrylic and methacrylic monomers and in both the presence and absence of light. These solutions yielded radicals, namely, nitroxides, only when containing methacrylic monomers, the presence of light having no effect. These observations support the "ene" addition/oxidation mechanism of nitroxide formation.

scholarly journals Accessing the distance range of interest in biomolecules: Site-directed spin labeling and DEER spectroscopy

2010 ◽  
Vol 24 (3-4) ◽  
pp. 283-288 ◽  
Author(s):  
Sabine Böhme ◽  
Heinz-Jürgen Steinhoff ◽  
Johann P. Klare
Keyword(s):  
Conformational Changes ◽  
Protein Interactions ◽  
Spin Labeling ◽  
Paramagnetic Resonance ◽  
Topological Information ◽  
Pulsed Epr ◽  
Site Directed Spin Labeling ◽  
Intermolecular Distances ◽  
And Function ◽  
Electron Paramagnetic

Investigations on the structure and function of biomolecules often depend on the availability of topological information to build up structural models or to characterize conformational changes during function. Electron paramagnetic resonance (EPR) spectroscopy in combination with site–directed spin labeling (SDSL) allow to determine intra- and intermolecular distances in the range from 4–70 Å, covering the range of interest for biomolecules. The approach does not require crystalline samples and is well suited also for molecules exhibiting intrinsic flexibility. This article is intended to give an overview on pulsed EPR in conjunction with SDSL to study protein interactions as well as conformational changes, exemplified on the tRNA modifying enzyme MnmE.

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