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Genetically Encoded Spin Labels for In Vitro and In-Cell EPR Studies of Native Proteins

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2015

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https://doi.org/10.1016/bs.mie.2015.05.023
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QIN, Peter Z., ed. and others. Electron Paramagnetic Resonance Investigations of Biological Systems by Using Spin Labels, Spin Probes, and Intrinsic Metal Ions ; Part A. Amsterdam: Elsevier, 2015, pp. 483-502. Methods in Enzymology. 563. ISBN 978-0-12-802834-6. Available under: doi: 10.1016/bs.mie.2015.05.023

Zusammenfassung

Electron paramagnetic resonance (EPR) spectroscopy in combination with site-directed spin labeling (SDSL) is a powerful approach to study the structure, dynamics, and interactions of proteins. The genetic encoding of the noncanonical amino acid spin-labeled lysine 1 (SLK-1) eliminates the need for any chemical labeling steps in SDSL-EPR studies and enables the investigation of native, endogenous proteins with minimal structural perturbation, and without the need to create unique reactive sites for chemical labeling. We report detailed experimental procedures for the efficient synthesis of SLK-1, the expression and purification of SLK-1-containing proteins under conditions that ensure maximal integrity of the nitroxide radical moiety, and procedures for intramolecular EPR distance measurements in proteins by double electron-electron resonance.

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540 Chemie

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Noncanonical amino acids, Genetic encoding, EPR spectroscopy, Site-directed spin labeling, In-cell EPR, DEER, Thioredoxin

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ISO 690SCHMIDT, Moritz J., Artem FEDOSEEV, Daniel SUMMERER, Malte DRESCHER, 2015. Genetically Encoded Spin Labels for In Vitro and In-Cell EPR Studies of Native Proteins. In: QIN, Peter Z., ed. and others. Electron Paramagnetic Resonance Investigations of Biological Systems by Using Spin Labels, Spin Probes, and Intrinsic Metal Ions ; Part A. Amsterdam: Elsevier, 2015, pp. 483-502. Methods in Enzymology. 563. ISBN 978-0-12-802834-6. Available under: doi: 10.1016/bs.mie.2015.05.023
BibTex
@incollection{Schmidt2015Genet-33218,
  year={2015},
  doi={10.1016/bs.mie.2015.05.023},
  title={Genetically Encoded Spin Labels for In Vitro and In-Cell EPR Studies of Native Proteins},
  number={563},
  isbn={978-0-12-802834-6},
  publisher={Elsevier},
  address={Amsterdam},
  series={Methods in Enzymology},
  booktitle={Electron Paramagnetic Resonance Investigations of Biological Systems by Using Spin Labels, Spin Probes, and Intrinsic Metal Ions ; Part A},
  pages={483--502},
  editor={Qin, Peter Z.},
  author={Schmidt, Moritz J. and Fedoseev, Artem and Summerer, Daniel and Drescher, Malte}
}
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