Rapid Analysis of DEER Signals Including Short Distances.

Double electron electron resonance (DEER) spectroscopy is an important technique to measure distance distributions () for studying protein structures and protein-protein interactions. DEER data analysis can at times become challenging due to the lack of a detailed analytical signal expression or numerical methods with rapid computation time. We have derived an analytical expression κ, which includes both the pseudo-secular dipolar coupling (PSDC) and the finite pulse effects, especially important for shorter distances.

MRI Denoising Using Pixel-Wise Threshold Selection.

Magnetic resonance imaging (MRI) has emerged as a promising technique for non-invasive medical imaging. The primary challenge in MRI is the trade-off between image visual quality and acquisition time. Current MRI image denoising algorithms employ global thresholding to denoise the whole image, which leads to inadequate denoising or image distortion.

Optimal Wavelet Selection for Signal Denoising.

Wavelet denoising plays a key role in removing noise from signals and is widely used in many applications. In denoising, selection of the mother wavelet is desirable for maximizing the separation of noise and signal coefficients in the wavelet domain for effective noise thresholding. At present, wavelet selection is carried out in a heuristic manner or using a trial-and-error that is time consuming and prone to error, including human bias.

Novel requirements for HAP2/GCS1-mediated gamete fusion in .

The ancestral gamete fusion protein, HAP2/GCS1, plays an essential role in fertilization in a broad range of taxa. To identify factors that may regulate HAP2/GCS1 activity, we screened mutants of the ciliate for behaviors that mimic knockout phenotypes in this species. Using this approach, we identified two new genes, and , whose products are necessary for membrane pore formation following mating type recognition and adherence.

An analysis of double-quantum coherence ESR in an N-spin system: Analytical expressions and predictions.

Electron spin resonance pulsed dipolar spectroscopy (PDS) has become popular in protein 3D structure analysis. PDS studies yield distance distributions between a pair or multiple pairs of spin probes attached to protein molecules, which can be used directly in structural studies or as constraints in theoretical predictions. Double-quantum coherence (DQC) is a highly sensitive and accurate PDS technique to study protein structures in the solid state and under physiologically relevant conditions.

Differentiating Unimodal and Multimodal Distributions in Pulsed Dipolar Spectroscopy Using Wavelet Transforms.

Site directed spin labeling has enabled protein structure determination using electron spin resonance (ESR) pulsed dipolar spectroscopy (PDS). Small details in a distance distribution can be key to understanding important protein structure-function relationships. A major challenge has been to differentiate unimodal and overlapped multimodal distance distributions.

Thermal Degradation of Thaumatin at Low pH and Its Prevention Using Alkyl Gallates.

Thaumatin, a potent sweet tasting protein extracted from the Katemfe Plant, is emerging as a natural alternative to synthetic non-nutritive sweeteners and flavor enhancer. As a food additive, its stability within the food matrix during thermal processing is of great interest to the food industry. When heated under neutral or basic conditions, thaumatin was found to lose its sweetness due to protein aggregation caused by sulfhydryl catalyzed disulfide bond interchange.

Structural insights into perilipin 3 membrane association in response to diacylglycerol accumulation.

Lipid droplets (LDs) are dynamic organelles that contain an oil core mainly composed of triglycerides (TAG) that is surrounded by a phospholipid monolayer and LD-associated proteins called perilipins (PLINs). During LD biogenesis, perilipin 3 (PLIN3) is recruited to nascent LDs as they emerge from the endoplasmic reticulum. Here, we analyze how lipid composition affects PLIN3 recruitment to membrane bilayers and LDs, and the structural changes that occur upon membrane binding.

Lewis acid-assisted reduction of nitrite to nitric and nitrous oxides via the elusive nitrite radical dianion.

Reduction of nitrite anions (NO) to nitric oxide (NO), nitrous oxide (NO) and ultimately dinitrogen (N) takes place in a variety of environments, including in the soil as part of the biogeochemical nitrogen cycle and in acidified nuclear waste. Nitrite reduction typically takes place within the coordination sphere of a redox-active transition metal. Here we show that Lewis acid coordination can substantially modify the reduction potential of this polyoxoanion to allow for its reduction under non-aqueous conditions (-0.

Membrane Binding Induces Distinct Structural Signatures in the Mouse Complexin-1C-Terminal Domain.

Complexins play a critical role in regulating SNARE-mediated exocytosis of synaptic vesicles. Evolutionary divergences in complexin function have complicated our understanding of the role these proteins play in inhibiting the spontaneous fusion of vesicles. Previous structural and functional characterizations of worm and mouse complexins have indicated the membrane curvature-sensing C-terminal domain of these proteins is responsible for differences in inhibitory function.

Theory and Least Squares Fitting of CW ESR Saturation Spectra Using the MOMD Model.

CW saturation experiments are widely used in ESR studies of relaxation processes in proteins and lipids. We develop the theory of saturation in ESR spectra in terms of its close relation with that of 2D-ELDOR. Our treatment of saturation is then based on the microscopic order macroscopic disorder (MOMD) model and can be used to fit the full CW saturation spectrum, rather than fitting just the peak-peak amplitude as a function of microwave field as is commonly done.

Structural Dynamics by NMR in the Solid State: II. The MOMD Perspective of the Dynamic Structure of Metal-Organic Frameworks Comprising Several Mobile Components.

We describe the application of the microscopic-order-macroscopic-disorder (MOMD) approach, developed for the analysis of dynamic H NMR lineshapes in the solid state, to unravel interactions among the constituents of metal-organic frameworks (MOFs) that comprise mobile components. MOMD was applied recently to University of Windsor Dynamic Material (UWDM) MOFs with one mobile crown ether per cavity. In this work, we study UWDM-9-, which comprises a mobile H-labeled phenyl-ring residue along with an isotopically unlabeled 24C8 crown ether.

The N-Terminal Domain of Aβ-Amyloid Fibril: The MOMD Perspective of its Dynamic Structure from NMR Lineshape Analysis.

We have developed the stochastic microscopic-order-macroscopic-disorder (MOMD) approach for elucidating dynamic structures in the solid-state from H NMR lineshapes. In MOMD, the probe experiences an effective/collective motional mode. The latter is described by a potential, , which represents the local spatial-restrictions, a local-motional diffusion tensor, , and key features of local geometry.

Negatively charged residues in the membrane ordering activity of SARS-CoV-1 and -2 fusion peptides.

Entry of coronaviruses into host cells is mediated by the viral spike protein. Previously, we identified the bona fide fusion peptides (FPs) for severe acute respiratory syndrome coronavirus ("SARS-1") and severe acute respiratory syndrome coronavirus-2 ("SARS-2") using electron spin resonance spectroscopy. We also found that their FPs induce membrane ordering in a Ca-dependent fashion.

Critical Negatively Charged Residues Are Important for the Activity of SARS-CoV-1 and SARS-CoV-2 Fusion Peptides.

Coronaviruses are a major infectious disease threat, and include the human pathogens of zoonotic origin SARS-CoV ("SARS-1"), SARS-CoV-2 ("SARS-2") and MERS-CoV ("MERS"). Entry of coronaviruses into host cells is mediated by the viral spike (S) protein. Previously, we identified that the domain immediately downstream of the S2' cleavage site is the FP (amino acids 798-835) for SARS-1 using ESR spectroscopy technology.

Benchmark Test and Guidelines for DEER/PELDOR Experiments on Nitroxide-Labeled Biomolecules.

Distance distribution information obtained by pulsed dipolar EPR spectroscopy provides an important contribution to many studies in structural biology. Increasingly, such information is used in integrative structural modeling, where it delivers unique restraints on the width of conformational ensembles. In order to ensure reliability of the structural models and of biological conclusions, we herein define quality standards for sample preparation and characterization, for measurements of distributed dipole-dipole couplings between paramagnetic labels, for conversion of the primary time-domain data into distance distributions, for interpreting these distributions, and for reporting results.

Highly Basic Clusters in the Herpes Simplex Virus 1 Nuclear Egress Complex Drive Membrane Budding by Inducing Lipid Ordering.

During replication of herpesviruses, capsids escape from the nucleus into the cytoplasm by budding at the inner nuclear membrane. This unusual process is mediated by the viral nuclear egress complex (NEC) that deforms the membrane around the capsid by oligomerizing into a hexagonal, membrane-bound scaffold. Here, we found that highly basic membrane-proximal regions (MPRs) of the NEC alter lipid order by inserting into the lipid headgroups and promote negative Gaussian curvature.

Dph3 Enables Aerobic Diphthamide Biosynthesis by Donating One Iron Atom to Transform a [3Fe-4S] to a [4Fe-4S] Cluster in Dph1-Dph2.

All radical -adenosylmethionine (radical-SAM) enzymes, including the noncanonical radical-SAM enzyme diphthamide biosynthetic enzyme Dph1-Dph2, require at least one [4Fe-4S](Cys) cluster for activity. It is well-known in the radical-SAM enzyme community that the [4Fe-4S](Cys) cluster is extremely air-sensitive and requires strict anaerobic conditions to reconstitute activity in vitro. Thus, how such enzymes function in vivo in the presence of oxygen in aerobic organisms is an interesting question.

Extraction of Weak Spectroscopic Signals with High Fidelity: Examples from ESR.

Noise impedes experimental studies by reducing signal resolution and/or suppressing weak signals. Signal averaging and filtering are the primary methods used to reduce noise, but they have limited effectiveness and lack capabilities to recover signals at low signal-to-noise ratios (SNRs). We utilize a wavelet transform-based approach to effectively remove noise from spectroscopic data.

SARS-CoV-2 Fusion Peptide has a Greater Membrane Perturbating Effect than SARS-CoV with Highly Specific Dependence on Ca.

Coronaviruses are a major infectious disease threat, and include the zoonotic-origin human pathogens SARS-CoV-2, SARS-CoV, and MERS-CoV (SARS-2, SARS-1, and MERS). Entry of coronaviruses into host cells is mediated by the spike (S) protein. In our previous ESR studies, the local membrane ordering effect of the fusion peptide (FP) of various viral glycoproteins including the S of SARS-1 and MERS has been consistently observed.

Microsecond dynamics in proteins by two-dimensional ESR. II. Addressing computational challenges.

Two-dimensional electron-electron double resonance (2D-ELDOR) provides extensive insight into molecular motions. Recent developments permitting experiments at higher frequencies (95 GHz) provide molecular orientational resolution, enabling a clearer description of the nature of the motions. In previous work, we provided simulations for the case of domain motions within proteins that are themselves slowly tumbling in a solution.

Microsecond Exchange Processes Studied by Two-Dimensional ESR at 95 GHz.

Exchange processes which include conformational change, protonation/deprotonation, and binding equilibria are routinely studied by 2D exchange NMR techniques, where information about the exchange of nuclei between environments with different NMR shifts is obtained from the development of cross-peaks. Whereas 2D NMR enables the real time study of millisecond and slower exchange processes, 2D ESR in the form of 2D-ELDOR (two-dimensional electron-electron double resonance) has the potential for such studies over the nanosecond to microsecond real time scales. Cross-peak development due to chemical exchange has been seen previously for semiquinones in ESR, but this is not possible for most common ESR probes, such as nitroxides, studied at typical ESR frequencies because, unlike NMR, the exchanging states yield ESR signals that are not resolved from each other within their respective line widths.

Engineered chemotaxis core signaling units indicate a constrained kinase-off state.

Bacterial chemoreceptors, the histidine kinase CheA, and the coupling protein CheW form transmembrane molecular arrays with remarkable sensing properties. The receptors inhibit or stimulate CheA kinase activity depending on the presence of attractants or repellants, respectively. We engineered chemoreceptor cytoplasmic regions to assume a trimer of receptor dimers configuration that formed well-defined complexes with CheA and CheW and promoted a CheA kinase-off state.