2013•Current Protocols in Protein ScienceOpen access

Pulsed EPR Distance Measurements in Soluble Proteins by Site‐Directed Spin Labeling (SDSL)

Ian Mitchelle S. de Vera, Mandy E. Blackburn, Luis Galiano, Gail E. Fanucci

Abstract

The resurgence of pulsed electron paramagnetic resonance (EPR) in structural biology centers on recent improvements in distance measurements using the double electron-electron resonance (DEER) technique. This unit focuses on EPR-based distance measurements by site-directed spin labeling (SDSL) of engineered cysteine residues in soluble proteins, with HIV-1 protease used as a model. To elucidate conformational changes in proteins, experimental protocols were optimized and existing data analysis programs were employed to derive distance-distribution profiles. Experimental considerations, sample preparation, and error analysis for artifact suppression are also outlined herein.

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Available abstract

Key concepts: Site-directed spin labeling, Electron paramagnetic resonance, Spin label, Chemistry, Nuclear magnetic resonance, Biophysics, Physics, Biology