Electron Paramagnetic Resonance Investigations of Biological Systems by Using Spin Labels, Spin Probes, and Intrinsic Metal Ions Part B (Volume 564) (Methods in Enzymology, Volume 564)

Experimental techniques capable of determining the structure and dynamics of proteins are continuously being developed in order to understand protein function. Among existing methods, site-directed spin labeling in combination with saturation recovery (SR) electron paramagnetic resonance spectroscopy contributes uniquely to the determination of secondary and tertiary protein structure under physiological conditions, independent of molecular weight and complexity. In addition, SR of spin labeled proteins was recently demonstrated to be sensitive to conformational exchange events with characteristic lifetimes on the order of μs, a time domain that presents a significant challenge to other spectroscopic techniques. In this chapter, we present the theoretical background necessary to understand the capabilities of SR as applied to spin labeled proteins, the instrumental requirements, and practical experimental considerations necessary to obtain interpretable data, and the use of SR to obtain information on protein: (1) secondary structure via solvent accessibility measurements, (2) tertiary structure using interspin distance measurements, and (3) conformational exchange. Content: Saturation recovery EPR and nitroxide spin labeling for exploring structure and dynamics in proteins -- High-pressure EPR and site-directed spin labeling for mapping molecular flexibility in proteins -- Exploring structure, dynamics, and topology of nitroxide spin-labeled proteins using continuous-wave electron paramagnetic resonance spectroscopy -- Bifunctional spin labeling of muscle proteins: accurate rotational dynamics, orientation, and distance by EPR -- EPR distance measurements in deuterated proteins -- Spin labeling and double electron-electron resonance (DEER) to deconstruct conformational ensembles of HIV protease -- Ionizable nitroxides for studying local electrostatic properties of lipid bilayers and protein systems by EPR -- Peptide-membrane interactions by spin-labeling EPR -- Structural characterization of membrane-curving proteins: site-directed spin labeling, EPR, and computational refinement -- Determining the secondary structure of membrane proteins and peptides via electron spin echo envelope modulation (ESEEM) spectroscopy -- Spin labeling studies of transmembranes signaling and transport: applications to phototaxis, ABC transporters and symporters -- Navigating membrane protein structure, dynamics, and energy landscapes using spin labeling and EPR spectroscopy -- Spin labeling of potassium channels -- Advanced EPR methods for studying conformational dynamics of nucleic acids -- An integrated spin-labeling/computational-modeling approach for mapping global structures of nucleic acids -- Overhauser dynamic nuclear polarization studies on local water dynamics -- Practical aspects of paramagnetic relaxation enhancement in biological macromolecules -- In vivo pO2 imaging of tumors: oxymetry with very low-frequency electron paramagnetic resonance -- Direct and repeated mesurement of heart and brain oxygenation using in vivo EPR oximetry -- Free radical imaging using in vivo dynamic nuclear polarization-MRI.

Electron Paramagnetic Resonance Investigations of Biological Systems by Using Spin Labels, Spin Probes, and Intrinsic Metal Ions Part A & B, are the latest volumes in the Methods in Enzymology series, continuing the legacy of this premier serial with quality chapters authored by leaders in the field. This volume covers research methods centered on the use of Electron Paramagnetic Resonance (EPR) techniques to study biological structure and function.

• Timely contribution that describes a rapidly changing field
• Leading researchers in the field
• Broad coverage: Instrumentation, basic theory, data analysis, and applications

Electron Paramagnetic Resonance Investigations of Biological Systems by Using Spin Labels, Spin Probes, and Intrinsic Metal Ions Part A & B , are the latest volumes in the Methods in Enzymology series, continuing the legacy of this premier serial with quality chapters authored by leaders in the field. This volume covers research methods centered on the use of Electron Paramagnetic Resonance (EPR) techniques to study biological structure and function. Timely contribution that describes a rapidly changing field Leading researchers in the field Broad coverage: Instrumentation, basic theory, data analysis, and applications