Structural Insights into the Dynamic Process of β2-Adrenergic Receptor Signaling
Cell 162(6):1431 (2015)
Structural Insights into the Dynamic Process of β2-Adrenergic Receptor Signaling.
Cell 161(5):1101 (2015)
G-protein-coupled receptors (GPCRs) transduce signals from the extracellular environment to intracellular proteins. To gain structural insight into the regulation of receptor cytoplasmic conformations by extracellular ligands during signaling, we examine the structural dynamics of the cytoplasmi...
Exploring Structure, Dynamics, and Topology of Nitroxide Spin-Labeled Proteins Using Continuous-Wave Electron Paramagnetic Resonance Spectroscopy.
Methods in Enzymology 564:59 (2015)
Structural and dynamical characterization of proteins is of central importance in understanding the mechanisms underlying their biological functions. Site-directed spin labeling (SDSL) combined with continuous-wave electron paramagnetic resonance (CW EPR) spectroscopy has shown the capability of...
Saturation Recovery EPR and Nitroxide Spin Labeling for Exploring Structure and Dynamics in Proteins.
Methods in Enzymology 564:3 (2015)
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 spectro...
High-Pressure EPR and Site-Directed Spin Labeling for Mapping Molecular Flexibility in Proteins.
Methods in Enzymology 564:29 (2015)
High hydrostatic pressure is a powerful probe of protein conformational flexibility. Pressurization reveals regions of elevated compressibility, and thus flexibility, within individual conformational states, but also shifts conformational equilibria such that "invisible" excited states become ac...
Differential dynamics of extracellular and cytoplasmic domains in denatured States of rhodopsin.
Biochemistry (Washington) 53(46):7160 (2014)
Rhodopsin is a model system for understanding membrane protein folding. Recently, conditions that allow maximally denaturing rhodopsin without causing aggregation have been determined, opening the door to the first structural characterization of denatured states of rhodopsin by nuclear magnetic ...
High resolution structure and double electron-electron resonance of the zebrafish voltage-dependent anion channel 2 reveal an oligomeric population.
Journal of Biological Chemistry 289(18):12566 (2014)
In recent years, there has been a vast increase in structural and functional understanding of VDAC1, but VDAC2 and -3 have been understudied despite having many unique phenotypes. One reason for the paucity of structural and biochemical characterization of the VDAC2 and -3 isoforms stems from th...
Self-association of arrestin family members.
Handbook of experimental pharmacology 219:205 (2014)
Mammals express four arrestin subtypes, three of which have been shown to self-associate. Cone photoreceptor-specific arrestin-4 is the only one that is a constitutive monomer. Visual arrestin-1 forms tetramers both in crystal and in solution, but the shape of its physiologically relevant soluti...
Rapid degeneration of rod photoreceptors expressing self-association-deficient arrestin-1 mutant.
Cellular Signalling 25(12):2613 (2013)
Arrestin-1 binds light-activated phosphorhodopsin and ensures timely signal shutoff. We show that high transgenic expression of an arrestin-1 mutant with enhanced rhodopsin binding and impaired oligomerization causes apoptotic rod death in mice. Dark rearing does not prevent mutant-induced cell ...
Conformational selection and adaptation to ligand binding in T4 lysozyme cavity mutants.
PNAS 110(46):E4306 (2013)
The studies presented here explore the relationship between protein packing and molecular flexibility using ligand-binding cavity mutants of T4 lysozyme. Although previously reported crystal structures of the mutants investigated show single conformations that are similar to the WT protein, site...
Technological advances in site-directed spin labeling of proteins.
Current Opinion in Structural Biology 23(5):725 (2013)
Molecular flexibility over a wide time range is of central importance to the function of many proteins, both soluble and membrane. Revealing the modes of flexibility, their amplitudes, and time scales under physiological conditions is the challenge for spectroscopic methods, one of which is site...
Structure and dynamics of an imidazoline nitroxide side chain with strongly hindered internal motion in proteins.
A disulfide-linked imidazoline nitroxide side chain (V1) has a similar and highly constrained internal motion at diverse topological sites in a protein, unlike that for the disulfide-linked pyrroline nitroxide side chain (R1) widely used in site directed spin labeling EPR. Crystal structures of ...
Engineering visual arrestin-1 with special functional characteristics.
Journal of Biological Chemistry 288(5):3394 (2013)
Arrestin-1 preferentially binds active phosphorylated rhodopsin. Previously, a mutant with enhanced binding to unphosphorylated active rhodopsin (Rh*) was shown to partially compensate for lack of rhodopsin phosphorylation in vivo. Here we showed that reengineering of the receptor binding surfac...
Structural states and dynamics of the D-loop in actin.
Biophysical Journal 103(5):930 (2012)
Conformational changes induced by ATP hydrolysis on actin are involved in the regulation of complex actin networks. Previous structural and biochemical data implicate the DNase I binding loop (D-loop) of actin in such nucleotide-dependent changes. Here, we investigated the structural and conform...
Structural States and Dynamics of the D-Loop in Actin
Biophysical Journal 103(5):930 (2012)
Conformational changes induced by ATP hydrolysis on actin are involved in the regulation of complex actin networks. Previous structural and biochemical data implicate the DNase I binding loop (D-loop) of actin in such nucleotide-dependent changes. Here, we investigated the structural a...
Measuring Protein Conformational Exchange Rates with Pressure-Jump Site Directed Spin Labeling EPR Spectroscopy
Biophysical Journal 102(3):405a (2012)
Site-directed spin labeling electron paramagnetic resonance study of the ORF1 protein from a mouse L1 retrotransposon.
Protein Science 20(7):1231 (2011)
Long interspersed nuclear element-1 is a highly abundant mammalian retrotransposon that comprises 17% of the human genome. L1 retrotransposition requires the protein encoded by open reading frame-1 (ORF1p), which binds single-stranded RNA with high affinity and functions as a nucleic acid chaper...
Site-directed spin labeling of a genetically encoded unnatural amino acid.
PNAS 106(51):21637 (2009)
The traditional site-directed spin labeling (SDSL) method, which utilizes cysteine residues and sulfhydryl-reactive nitroxide reagents, can be challenging for proteins that contain functionally important native cysteine residues or disulfide bonds. To make SDSL amenable to any protein, we introd...
Effects of binding factors on structural elements in F-actin.
Biochemistry (Washington) 48(2):370 (2009)
Understanding the dynamics of the actin filament is essential to a detailed description of their interactions and role in the cell. Previous studies have linked the dynamic properties of actin filaments (F-actin) to three structural elements contributing to a hydrophobic pocket, namely, the hydr...
High-resolution distance mapping in rhodopsin reveals the pattern of helix movement due to activation.
PNAS 105(21):7439 (2008)
Site-directed spin labeling has qualitatively shown that a key event during activation of rhodopsin is a rigid-body movement of transmembrane helix 6 (TM6) at the cytoplasmic surface of the molecule. To place this result on a quantitative footing, and to identify movements of other helices upon ...