Combining site-directed spin labeling in vivo and in-cell EPR distance determination

Literature Information

Publication Date 2020-02-18
DOI 10.1039/C9CP05584C
Impact Factor 3.676
Authors

Pia Widder, Julian Schuck, Daniel Summerer, Malte Drescher


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Abstract

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.

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Source Journal

Physical Chemistry Chemical Physics

Physical Chemistry Chemical Physics
CiteScore: 5.5
Self-citation Rate: 10.3%
Articles per Year: 3036

Physical Chemistry Chemical Physics (PCCP) is an international journal co-owned by 19 physical chemistry and physics societies from around the world. This journal publishes original, cutting-edge research in physical chemistry, chemical physics and biophysical chemistry. To be suitable for publication in PCCP, articles must include significant innovation and/or insight into physical chemistry; this is the most important criterion that reviewers and Editors will judge against when evaluating submissions. The journal has a broad scope and welcomes contributions spanning experiment, theory, computation and data science. Topical coverage includes spectroscopy, dynamics, kinetics, statistical mechanics, thermodynamics, electrochemistry, catalysis, surface science, quantum mechanics, quantum computing and machine learning. Interdisciplinary research areas such as polymers and soft matter, materials, nanoscience, energy, surfaces/interfaces, and biophysical chemistry are welcomed if they demonstrate significant innovation and/or insight into physical chemistry. Joined experimental/theoretical studies are particularly appreciated when complementary and based on up-to-date approaches.

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