Versatile Trityl Spin Labels for Nanometer Distance Measurements on Biomolecules In Vitro and within Cells

2016 ◽  
Vol 129 (1) ◽  
pp. 183-187 ◽  
Author(s):  
J. Jacques Jassoy ◽  
Andreas Berndhäuser ◽  
Fraser Duthie ◽  
Sebastian P. Kühn ◽  
Gregor Hagelueken ◽  
...  
Keyword(s):  
Spin Labels ◽  
Distance Measurements

Versatile Trityl Spin Labels for Nanometer Distance Measurements on Biomolecules In Vitro and within Cells

2016 ◽  
Vol 56 (1) ◽  
pp. 177-181 ◽  
Author(s):  
J. Jacques Jassoy ◽  
Andreas Berndhäuser ◽  
Fraser Duthie ◽  
Sebastian P. Kühn ◽  
Gregor Hagelueken ◽  
...  
Keyword(s):  
Spin Labels ◽  
Distance Measurements

Conformational changes of the chaperone SecB upon binding to a model substrate – bovine pancreatic trypsin inhibitor (BPTI)

2011 ◽  
Vol 392 (10) ◽  
pp. 849-858 ◽  
Author(s):  
Michaela M. Haimann ◽  
Yasar Akdogan ◽  
Reinhard Philipp ◽  
Raghavan Varadarajan ◽  
Dariush Hinderberger ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Trypsin Inhibitor ◽  
Tertiary Structure ◽  
Substrate Binding ◽  
Spin Labeling ◽  
Bovine Pancreatic Trypsin Inhibitor ◽  
Spin Labels ◽  
Distance Measurements ◽  
Pancreatic Trypsin Inhibitor

Abstract SecB is a homotetrameric cytosolic chaperone that forms part of the protein translocation machinery in E. coli. Due to SecB, nascent polypeptides are maintained in an unfolded translocation-competent state devoid of tertiary structure and thus are guided to the translocon. In vitro SecB rapidly binds to a variety of ligands in a non-native state. We have previously investigated the bound state conformation of the model substrate bovine pancreatic trypsin inhibitor (BPTI) as well as the conformation of SecB itself by using proximity relationships based on site-directed spin labeling and pyrene fluorescence methods. It was shown that SecB undergoes a conformational change during the process of substrate binding. Here, we generated SecB mutants containing but a single cysteine per subunit or an exposed highly reactive new cysteine after removal of the nearby intrinsic cysteines. Quantitative spin labeling was achieved with the methanethiosulfonate spin label (MTS) at positions C97 or E90C, respectively. Highfield (W-band) electron paramagnetic resonance (EPR) measurements revealed that with BPTI present the spin labels are exposed to a more polar/hydrophilic environment. Nanoscale distance measurements with double electron-electron resonance (DEER) were in excellent agreement with distances obtained by molecular modeling. Binding of BPTI also led to a slight change in distances between labels at C97 but not at E90C. While the shorter distance in the tetramer increased, the larger diagonal distance decreased. These findings can be explained by a widening of the tetrameric structure upon substrate binding much like the opening of two pairs of scissors.

Triplet Fullerenes as Prospective Spin Labels for Nanoscale Distance Measurements by Pulsed Dipolar EPR Spectroscopy

2019 ◽  
Vol 131 (38) ◽  
pp. 13405-13409 ◽  
Author(s):  
Olesya A. Krumkacheva ◽  
Ivan O. Timofeev ◽  
Larisa V. Politanskaya ◽  
Yuliya F. Polienko ◽  
Evgeny V. Tretyakov ◽  
...  
Keyword(s):  
Epr Spectroscopy ◽  
Spin Labels ◽  
Distance Measurements

scholarly journals Probing the solution structure of the E. coli multidrug transporter MdfA using DEER distance measurements with nitroxide and Gd(III) spin labels

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Eliane H. Yardeni ◽  
Thorsten Bahrenberg ◽  
Richard A. Stein ◽  
Smriti Mishra ◽  
Elia Zomot ◽  
...  
Keyword(s):  
Solution Structure ◽  
Spin Labels ◽  
Multidrug Transporter ◽  
Distance Measurements ◽  
E Coli

A new approach to distance measurements between two spin labels in the >10 nm range

2017 ◽  
Vol 19 (7) ◽  
pp. 5222-5229 ◽  
Author(s):  
A. Blank
Keyword(s):  
Esr Spectroscopy ◽  
Dipolar Interaction ◽  
Spin Labels ◽  
Interaction Frequency ◽  
Distance Measurements ◽  
New Approach ◽  
Nm Range

ESR spectroscopy can be efficiently used to acquire the distance between two spin labels placed on a macromolecule by measuring their mutual dipolar interaction frequency, as long as the distance is not greater than ∼10 nm.

Pulsed electron–electron double resonance: beyond nanometre distance measurements on biomacromolecules

2011 ◽  
Vol 434 (3) ◽  
pp. 353-363 ◽  
Author(s):  
Gunnar W. Reginsson ◽  
Olav Schiemann
Keyword(s):  
Protein Complexes ◽  
Double Resonance ◽  
Resonance Method ◽  
Spin Labels ◽  
Coupling Mechanism ◽  
Distance Measurements ◽  
Paramagnetic Resonance ◽  
Distance Distributions ◽  
Nitroxide Spin Labels ◽  
Amino Acid Radicals

PELDOR (or DEER; pulsed electron–electron double resonance) is an EPR (electron paramagnetic resonance) method that measures via the dipolar electron–electron coupling distances in the nanometre range, currently 1.5–8 nm, with high precision and reliability. Depending on the quality of the data, the error can be as small as 0.1 nm. Beyond mere mean distances, PELDOR yields distance distributions, which provide access to conformational distributions and dynamics. It can also be used to count the number of monomers in a complex and allows determination of the orientations of spin centres with respect to each other. If, in addition to the dipolar through-space coupling, a through-bond exchange coupling mechanism contributes to the overall coupling both mechanisms can be separated and quantified. Over the last 10 years PELDOR has emerged as a powerful new biophysical method without size restriction to the biomolecule to be studied, and has been applied to a large variety of nucleic acids as well as proteins and protein complexes in solution or within membranes. Small nitroxide spin labels, paramagnetic metal ions, amino acid radicals or intrinsic clusters and cofactor radicals have been used as spin centres.

scholarly journals Trueness and Precision of Three-Dimensional Digitizing Intraoral Devices

2018 ◽  
Vol 2018 ◽  
pp. 1-10 ◽  
Author(s):  
Hussam Mutwalli ◽  
Michael Braian ◽  
Deyar Mahmood ◽  
Christel Larsson
Keyword(s):  
Three Dimensional ◽  
Coordinate Measuring Machine ◽  
In Vitro Study ◽  
Vitro Study ◽  
Distance Measurements ◽  
True Value ◽  
Measuring Machine ◽  
One Way Anova ◽  
Intraoral Devices

Aim. To measure the trueness and precision under repeatable conditions for different intraoral scanners (IOSs) when scanning fully edentulous arch with multiple implants. Materials and Methods. Three IOSs and one industrial scanner were used to scan one edentulous master cast containing five implant scan bodies and three spheres. The cast was scanned thirty times with each scanner device. All scans were analyzed in the inspect software, and three-dimensional locations of the implants and the interarch distance between the spheres were measured. The values were compared to measurements made with one coordinate measuring machine (true value). One-way ANOVA was used to calculate the differences between IOSs and in comparison with the true value. Results. Significant differences were found between all IOSs. For the implant measurements, Trios 3 had the lowest trueness (≤114 μm), followed by Trios 3 mono (≤63 μm) and Itero element (≤−41 μm). Trios had the lowest precision (≤135 μm), followed by Itero element (≤101 μm) and Trios 3 mono (≤100 μm). With regard to the interarch distance measurements, Trios 3 had the lowest trueness (≤68 μm), followed by Trios 3 mono (≤45 μm) and Itero element (≤40 μm). Trios 3 had the lowest precision (≤206 μm), followed by Itero element (≤124 μm) and Trios 3 mono (≤111 μm). Conclusion. The results from this in vitro study suggest that precision is low for the tested IOS devices when scanning fully edentulous arches with multiple implants.

Generic tags for Mn(ii) and Gd(iii) spin labels for distance measurements in proteins

2017 ◽  
Vol 19 (39) ◽  
pp. 26944-26956 ◽  
Author(s):  
Yin Yang ◽  
Yan-Jun Gong ◽  
Aleksei Litvinov ◽  
Hong-Kai Liu ◽  
Feng Yang ◽  
...  
Keyword(s):  
Spin Label ◽  
Metal Ion ◽  
Coordination Mode ◽  
Distance Distribution ◽  
Spin Labels ◽  
Distance Measurements

The coordination mode of the metal ion in the spin label affects the distance distribution determined by DEER distance measurements.

scholarly journals Combining Site-Directed Spin Labeling in Vivo and In-Cell EPR Distance Determination

2019 ◽  
Author(s):  
Pia Widder ◽  
Julian Schuck ◽  
Daniel Summerer ◽  
Malte Drescher
Keyword(s):  
Spin Labeling ◽  
Distance Measurements ◽  
Paramagnetic Resonance ◽  
Distance Determination ◽  
Bioorthogonal Labeling ◽  
In Vitro Measurements ◽  
Site Directed Spin Labeling ◽  
Double Electron Electron Resonance

Structural studies on proteins directly in their native environment are required for a comprehensive understanding of their function. Electron paramagnetic resonance (EPR) spectroscopy and in particular double electron-electron resonance (DEER) distance determination are suited to investigate spin-labeled proteins directly in the cell. The combination of intracellular bioorthogonal labeling with in-cell DEER measurements does not require additional purification or delivery steps of spin-labeled protein to the cells. In this study, we express eGFP in E.coli and use copper-catalyzed azide-alkyne cycloaddition (CuAAC) for the site-directed spin labeling of the protein in vivo, followed by in-cell EPR distance determination. Inter-spin distance measurements of spin-labeled eGFP agree with in vitro measurements and calculations based on the rotamer library of the spin label.<br>

scholarly journals Combining Site-Directed Spin Labeling in Vivo and In-Cell EPR Distance Determination

2019 ◽  
Author(s):  
Pia Widder ◽  
Julian Schuck ◽  
Daniel Summerer ◽  
Malte Drescher
Keyword(s):  
Spin Labeling ◽  
Distance Measurements ◽  
Paramagnetic Resonance ◽  
Distance Determination ◽  
Bioorthogonal Labeling ◽  
In Vitro Measurements ◽  
Site Directed Spin Labeling ◽  
Double Electron Electron Resonance

Structural studies on proteins directly in their native environment are required for a comprehensive understanding of their function. Electron paramagnetic resonance (EPR) spectroscopy and in particular double electron-electron resonance (DEER) distance determination are suited to investigate spin-labeled proteins directly in the cell. The combination of intracellular bioorthogonal labeling with in-cell DEER measurements does not require additional purification or delivery steps of spin-labeled protein to the cells. In this study, we express eGFP in E.coli and use copper-catalyzed azide-alkyne cycloaddition (CuAAC) for the site-directed spin labeling of the protein in vivo, followed by in-cell EPR distance determination. Inter-spin distance measurements of spin-labeled eGFP agree with in vitro measurements and calculations based on the rotamer library of the spin label.<br>

Sign in / Sign up

Export Citation Format

Share Document