A5081581715- Enzyme Structure and Function
- Protein Structure and Dynamics
- Photosynthetic Processes and Mechanisms
- Photoreceptor and optogenetics research
- Hemoglobin structure and function
- Mass Spectrometry Techniques and Applications
- Advanced Electron Microscopy Techniques and Applications
- Receptor Mechanisms and Signaling
- HIV Research and Treatment
- Advanced Fluorescence Microscopy Techniques
- Metal-Catalyzed Oxygenation Mechanisms
- Spectroscopy and Quantum Chemical Studies
- Metabolomics and Mass Spectrometry Studies
- Amino Acid Enzymes and Metabolism
- Advanced MRI Techniques and Applications
- Pancreatic function and diabetes
- HIV/AIDS drug development and treatment
- Advanced Proteomics Techniques and Applications
- Protein purification and stability
- X-ray Spectroscopy and Fluorescence Analysis
- Proteins in Food Systems
- RNA and protein synthesis mechanisms
- Atomic and Subatomic Physics Research
- RNA Interference and Gene Delivery
- Advanced biosensing and bioanalysis techniques
University of Chicago
2014-2024
Argonne National Laboratory
2014-2023
MAX IV Laboratory
2020
Lund University
2020
University of Gothenburg
2020
BioCARS, a NIH-supported national user facility for macromolecular time-resolved X-ray crystallography at the Advanced Photon Source (APS), has recently completed commissioning of an upgraded undulator-based beamline optimized single-shot laser-pump X-ray-probe measurements with time resolution as short 100 ps. The source consists two in-line undulators periods 23 and 27 mm that together provide high-flux pink-beam capability 12 keV well first-harmonic coverage from 6.8 to 19 keV. A...
A major goal in biomedical science is to move beyond static images of proteins and other biological macromolecules the internal dynamics underlying their function. This level study necessary understand how these molecules work engineer new functions modulators Stemming from a visionary commitment this problem by Keith Moffat decades ago, community structural biologists has now enabled set x-ray scattering technologies for observing intramolecular at atomic resolution over broad range...
Proteins serve as molecular machines in performing their biological functions, but the detailed structural transitions are difficult to observe native aqueous environments real time. For example, despite extensive studies, solution-phase structures of intermediates along allosteric pathways for between relaxed (R) and tense (T) forms have been elusive. In this work, we employed picosecond X-ray solution scattering novel analysis track details dynamics wild-type homodimeric hemoglobin (HbI)...
The HIV-1 Envelope (Env) glycoprotein facilitates host cell fusion through a complex series of receptor-induced structural changes. Although remarkable progress has been made in understanding the structures various Env conformations, microsecond timescale dynamics have not studied experimentally. Here, we used time-resolved, temperature-jump small-angle x-ray scattering to monitor rearrangements an SOSIP ectodomain construct with precision. In two distinct variants, detected transition that...
Biological functions frequently require protein-protein interactions that involve secondary and tertiary structural perturbation. Here we study dissociation reassociation dynamics in insulin, a model system for protein oligomerization. Insulin dimer into monomers was induced by nanosecond temperature-jump (T-jump) of ∼8 °C aqueous solution, the resulting solvent were tracked time-resolved X-ray solution scattering (TRXSS) on time scales 10 ns to 100 ms. The signals revealed formation five...
Understanding protein folding pathways is crucial to deciphering the principles of structure and function. Here, we investigate unfolding dynamics 35‐residue villin headpiece (HP35) a norleucine‐substituted variant (2F4K) using combination experimental computational techniques. Time‐resolved X‐ray solution scattering (TRXSS) coupled with equilibrium Molecular Dynamics (MD) simulations Markov State modeling reveals distinct mechanisms between two variants: HP35 2F4K. Specifically, exhibits...
Photosynthetic water oxidation is a fundamental process that sustains the biosphere. A Mn$_{4}$Ca cluster embedded in photosystem II protein environment responsible for production of atmospheric oxygen. Here, time-resolved x-ray emission spectroscopy (XES) was used to observe oxygen formation real time. These experiments reveal evolution step, initiated by three sequential laser flashes, accompanied rapid (within 50 $\mu$s) changes Mn K$\beta$ XES spectrum. However, no core above all...
Serial methods for crystallography have the potential to enable dynamic structural studies of protein targets that been resistant single-crystal strategies. The use serial data-collection strategies can circumvent challenges associated with radiation damage and repeated reaction initiation. This work utilizes a microfluidic crystallization platform time-resolved Laue diffraction analysis macroscopic crystals photoactive yellow (PYP). Reaction initiation was achieved via pulsed laser...
The protein folding process often proceeds through partially folded transient states. Therefore, a structural understanding of these disordered states is crucial for developing mechanistic models the process. Characterization unfolded remains challenging due to their nature, and incorporating multiple methods necessary. Combining time-resolved x-ray solution scattering (TRXSS) signal with molecular dynamics (MD), we are able characterize bovine α-lactalbumin, model system widely used...
Renewed interest in room-temperature diffraction has been prompted by the desire to observe structural dynamics of proteins as they function. Serial crystallography, an experimental strategy that aggregates small pieces data from a large uniform pool crystals, demonstrated at synchrotrons and X-ray free-electron lasers. This work utilizes microfluidic crystallization platform for serial Laue macroscopic crystals proposes collection slices many individual is realistic solution difficulties...
It is well-known that biological samples undergo X-ray-induced degradation. One of the fastest occurring processes involves redox modifications (reduction or oxidation) redox-active cofactors in proteins. Here we analyze room-temperature data on photoreduction Mn ions oxygen-evolving complex (OEC) photosystem II, one most radiation damage-sensitive proteins and a key constituent natural photosynthesis plants, green algae, cyanobacteria. Time-resolved X-ray emission spectroscopy with...
Structural changes in global conformation and chromophore of photoactive yellow protein are probed real time by time-resolved X-ray solution scattering transient absorption spectroscopy.
Abstract The Light-oxygen-voltage-sensing domain (LOV) superfamily, found in enzymes and signal transduction proteins, plays a crucial role converting light signals into structural signals, mediating various biological mechanisms. While time-resolved spectroscopic studies have revealed the dynamics of LOV-domain chromophore’s electronic structures, understanding changes protein moiety, particularly regarding light-induced dimerization, remains challenging. Here, we utilize X-ray...
Direct tracking of protein structural dynamics during folding–unfolding processes is important for understanding the roles hierarchic factors in formation functional proteins. Using cytochrome c (cyt c) as a platform, we investigated its folding triggered by local environmental changes (i.e., pH or heme iron center oxidation/spin/ligation states) with time-resolved X-ray solution scattering measurements. Starting from partially unfolded cyt c, sudden drop initiated light excitation photoacid...
Many biomaterials can adapt to changes in the local biological environment (such as pH, temperature, or ionic composition) order regulate function deliver a payload. Such adaptation environmental perturbation is typically hierarchical process that begins with response at structural level and then propagates supramolecular macromolecular scales. Understanding fast dynamics occur upon important for rational design of functional biomaterials. However, few nanosecond time-resolved methods probe...
In the past few decades, prediction of macromolecular structures beyond native conformation has been aided by development molecular dynamics (MD) protocols aimed at exploration energetic landscape proteins. Yet, computed do not always agree with experimental observables, calling for further MD strategies to bring computations and experiments closer together. Here, we report a scalable, efficient simulation approach that incorporates an x-ray solution scattering signal as driving force...
The paradigm of "detection-before-destruction" was tested for a metalloprotein complex exposed at room temperature to the high x-ray flux typical third generation synchrotron sources. Following progression induced damage by Mn Kβ emission spectroscopy, we demonstrated feasibility collecting data on electronic structure native Photosystem II, trans-membrane containing Mn(4)Ca cluster. determined non-damaging observation timeframe (about 100 milliseconds using continuous monochromatic beam,...
Parameters of local heme structure and overall conformation are tracked to reveal conformational influences on ligation states.
The correct folding of proteins is paramount importance for their function, and protein misfolding believed to be the primary cause a wide range diseases. Protein has been investigated with time-averaged methods time-resolved spectroscopy, but observing structural dynamics unfolding process in real-time challenging. Here, we demonstrate an approach directly reveal changes reaction. We use nano- millisecond x-ray solution scattering probe apomyoglobin. reaction was triggered using temperature...
The Trp-cage miniprotein is one of the smallest systems to exhibit a stable secondary structure and fast-folding dynamics, serving as an apt model system study transient intermediates with both experimental computational analyses. Previous spectroscopic characterizations that have been done on inferred single intermediate pathway from folded unfolded basins. We aim bridge understanding structural folding dynamics microsecond-time scales, by utilizing time-resolved X-ray solution scattering...
A fundamental problem in biological sciences is understanding how macromolecular machines work and the structural changes of a molecule are connected to its function. Time-resolved techniques vital this regard essential for dynamics biomolecules. small- wide-angle X-ray solution scattering has capability provide multitude information about kinetics global molecules under their physiological conditions. However, standard protocols such time-resolved measurements often require significant...
The fluorescent protein Dronpa undergoes reversible photoswitching reactions between the bright "on" and dark "off" states via photoisomerization proton transfer reactions. We report room temperature crystal structure of fast switching Met159Thr mutant at 2.0-Å resolution in on state. Structural differences with wild type include shifted backbone positions strand β8 containing Thr159 as well an altered A-C dimer interface involving strands β7, β8, β10, β11. mutation increases cavity volume...