CW ESR spectroscopy and protein spin labeling in membrane biology

Biological membranes define cellular and organelle boundaries, and perform vital functions, providing transport, recognition, signaling, and interaction with other cells. These membranes are majorly composed of lipid bilayers and membrane proteins. Membrane proteins perform most membrane functions. Based on their localization, they are classified as integral and peripheral proteins. In this overview, we provide basic information about membrane proteins structure, conformational dynamics, and functions, and outline the methodologies used to produce highly-pure functional membrane proteins forin vitrobiophysical characterizations based on selected examples. To this end, expression of membrane proteins in a host, their extraction, purification and reconstitution in model lipid bilayers are described. Further, biophysical approaches play key role in elucidation of the structure and function of membrane proteins. Our focus here is on the technique of continuous wave electron paramagnetic/spin resonance (CW ESR) spectroscopy applied to spin-labeled membrane proteins. We describe the basic principles of membrane proteins labeling with nitroxide spin labels (paramagnetic tags) and how the CW ESR can be successfully used in elucidating the conformational dynamics of such proteins. We describe the basic principles of the CW ESR technique. The capability of this technique to characterize physiologically relevant conformational dynamics of proteins is demonstrated using two examples of CW ESR studies on spin-labeled human Tau and influenza A M2 proteins. The method is highly suitable to study physiological structure-function relationships of a broad range of proteins, and to explain the malfunctional states of proteins linked to diseases. This review is directed to the broader biophysical community with interest in molecular biophysics of biological membranes.