A5006725012- Endoplasmic Reticulum Stress and Disease
- Receptor Mechanisms and Signaling
- Mass Spectrometry Techniques and Applications
- Advanced Proteomics Techniques and Applications
- Neuropeptides and Animal Physiology
- Diabetes Treatment and Management
- Monoclonal and Polyclonal Antibodies Research
- Click Chemistry and Applications
- Epigenetics and DNA Methylation
- Plant nutrient uptake and metabolism
- Enzyme Structure and Function
- Protein Degradation and Inhibitors
- Plant Stress Responses and Tolerance
- Protein Structure and Dynamics
- Computational Drug Discovery Methods
- Estrogen and related hormone effects
- Amino Acid Enzymes and Metabolism
- Neuroendocrine Tumor Research Advances
- Photoreceptor and optogenetics research
- thermodynamics and calorimetric analyses
- Signaling Pathways in Disease
- Biotin and Related Studies
- Ubiquitin and proteasome pathways
- Drug Transport and Resistance Mechanisms
- Chemokine receptors and signaling
Pfizer (United States)
2016-2024
Foton Motors (China)
2016-2020
Scripps Research Institute
2011-2018
Worldwide Clinical Trials (United States)
2016
IONICS Mass Spectrometry (Canada)
2013
Duke University
2008-2012
Environmental Protection Agency
2012
Duke University Hospital
2010
Duke Medical Center
2010
Cleveland Clinic Lerner College of Medicine
2009
Musical Chairs The plant hormone abscisic acid (ABA) helps plants to respond changes in the environment, such as drought. Physiological responses are initiated when ABA binds its receptor. In absence of ABA, downstream kinases held inactive by phosphatases. Soon et al. (p. 85 , published online 24 November; see Perspective Leung ) now show that both hormone-receptor complex and kinase bind same site on phosphatase. Thus, presence hormone, phosphatase is occupied unable interfere with activity.
Proteolysis targeting chimeras (PROTACs) are heterobifunctional small molecules that simultaneously bind to a target protein and an E3 ligase, thereby leading ubiquitination subsequent degradation of the target. They present exciting opportunity modulate proteins in manner independent enzymatic or signaling activity. As such, they have recently emerged as attractive mechanism explore previously "undruggable" targets. Despite this interest, fundamental questions remain regarding parameters...
Hydrogen/deuterium exchange mass spectrometry (HDX-MS) is an established method for the interrogation of protein conformation and dynamics. While data analysis challenge HDX-MS has been addressed by a number software packages, new computational tools are needed to keep pace with improved methods throughput this technique. To address these needs, we report integrated desktop program titled HDX Workbench, which facilitates automation, management, visualization, statistical cross-comparison...
Described here is a new technique, termed SPROX (stability of proteins from rates oxidation), that can be used to measure the thermodynamic stability and protein-ligand complexes. utilizes hydrogen peroxide in presence increasing concentrations chemical denaturant oxidize proteins. The extent oxidation at given time determined as function concentration using either electrospray or matrix-assisted laser desorption/ionization mass spectrometry. Ultimately, dependence reaction rate evaluate...
Knowledge about the protein targets of therapeutic agents is critical for understanding drug mode action. Described here a mass spectrometry-based proteomics method identifying target(s) molecules that potentially applicable to any compound. The method, which involves making thermodynamic measurements protein-folding reactions in complex biological mixtures detect protein–drug interactions, demonstrated an experiment identify yeast immunosuppressive drug, cyclosporin A (CsA). Two ten...
Abscisic acid (ABA) is an essential hormone that controls plant growth, development, and responses to abiotic stresses. Central for ABA signaling the ABA-mediated autoactivation of three monomeric Snf1-related kinases (SnRK2.2, -2.3, -2.6). In absence ABA, SnRK2s are kept in inactive state by forming physical complexes with type 2C protein phosphatases (PP2Cs). Upon relief this inhibition, SnRK2 can autoactivate through unknown mechanisms. Here, we report crystal structures full-length...
Class B G protein-coupled receptors are composed of an extracellular domain (ECD) and a seven-transmembrane (7TM) domain, their signalling is regulated by peptide hormones. Using hybrid structural biology approach together with the ECD 7TM crystal structures glucagon receptor (GCGR), we examine relationship between full-length conformation ligand binding. Molecular dynamics (MD) disulfide crosslinking studies suggest that apo-GCGR can adopt both open closed associated extensive contacts...
Electrophilic small molecules that can reversibly modify proteins are of growing interest in drug discovery. However, the ability to study reversible covalent probes live cells be limited by their reactivity after cell lysis and proteomic workflows, leading scrambling signal loss. We describe how thiomethyltetrazines function as warheads for cysteine modification, this dynamic labeling behavior "switched off" via bioorthogonal chemistry inside cells. Simultaneously, tetrazine serves a...
Inhibition of β-secretase BACE1 is considered one the most promising approaches for treating Alzheimer's disease. Several structurally distinct inhibitors have been withdrawn from development after inducing ocular toxicity in animal models, but target mediating this has not identified. Here we use a clickable photoaffinity probe to identify cathepsin D (CatD) as principal off-target human cells. We find that several blocked CatD activity cells with much greater potency than displayed...
Alanine-serine-cysteine transporter 2 (ASCT2, SLC1A5) is the primary of glutamine in cancer cells and regulates mTORC1 signaling pathway. The SLC1A5 function involves finely tuned orchestration two domain movements that include substrate-binding transport scaffold domain. Here, we present cryo-EM structures human its complex with substrate, L-glutamine an outward-facing conformation. These reveal insights into conformation critical ECL2a loop which connects domains, thus allowing rigid body...
Protein arginine methyltransferase 1 (PRMT1)-dependent methylation contributes to the onset and progression of numerous diseases (e.g., cancer, heart disease, ALS); however, regulatory mechanisms that control PRMT1 activity are relatively unexplored. We therefore set out decipher how phosphorylation regulates activity. Curated mass spectrometry data identified Tyr291, a residue adjacent conserved THW loop, as being phosphorylated. Natural unnatural amino acid mutagenesis, including...
The orphan nuclear receptor liver homolog 1 (LRH-1; NR5A2) is a potent regulator of cholesterol metabolism and bile acid homeostasis. Recently, LRH-1 has been shown to play an important role in intestinal inflammation the progression estrogen positive negative breast cancers pancreatic cancer. Structural studies have revealed that can bind phospholipids dietary phospholipid dilauroylphosphatidylcholine activates activity rodents. Here we characterize novel synthetic nonphospholipid small...
Knowledge about the structural and biophysical properties of proteins when they are free in solution and/or complexes with other molecules is essential for understanding biological processes that regulate. Such knowledge also important to drug discovery efforts, particularly those focused on development therapeutic agents protein targets. In last decade a variety different covalent labeling techniques have been used combination mass spectrometry probe solution-phase structures protein-ligand...
Current methods for the large-scale characterization of disease states generally rely on analysis gene and/or protein expression levels. These existing fail to detect proteins with disease-related functions and unaltered Here we describe use thermodynamic measurements folding stability states. Using Stable Isotope Labeling Amino Acids in Cell Culture Stability Proteins from Rates Oxidation (SILAC-SPROX) technique, assayed ∼800 changes three different cell culture models breast cancer...
Abstract Cystic Fibrosis (CF) is caused by mutations in the Transmembrane Conductance Regulator (CFTR). Mutations associated with CF cause loss-of-function CFTR leading to salt imbalance epithelial tissues. Kalydeco (also called VX-770 or ivacaftor) was approved for treatment 2012 but little known regarding compound’s interactions including site of binding mechanisms action. In this study we use hydrogen/deuterium exchange (HDX) coupled mass spectrometry assess conformational dynamics a...
Stability proteomics techniques that do not require drug modifications have emerged as an attractive alternative to affinity purification methods in target engagement studies. Two representative include the chemical-denaturation-based SPROX (Stability of Proteins from Rates Oxidation), which utilizes peptide-level quantification and thermal-denaturation-based TPP (Thermal Proteome Profiling), protein-level quantification. Recently, "OnePot" strategy was adapted for both increase throughput....
Described here is a mass spectrometry-based screening assay for the detection of protein-ligand binding interactions in multicomponent protein mixtures. The utilizes an oxidation labeling protocol that involves using hydrogen peroxide to selectively oxidize methionine residues proteins order probe solvent accessibility these as function temperature. extent which are oxidized after specified reaction times at range temperatures determined MALDI analysis intact and/or LC-MS tryptic peptide...
Plasma protein binding (PPB) studies on the SARS-CoV-2 main protease inhibitor nirmatrelvir revealed considerable species differences primarily in dog and rabbit, which prompted further investigations into biochemical basis for these differences.The unbound fraction (fu) of rabbit plasma was concentration (2–200 µM)-dependent (dog fu,p 0.024–0.69, 0.010–0.82). Concentration (0.1–100 serum albumin (SA) (fu,SA 0.040–0.82) alpha-1-acid glycoprotein (AAG) (fu,AAG 0.050–0.64) observed dogs....
Described here is a stable isotope labeling protocol that can be used with chemical modification- and mass spectrometry-based protein-ligand binding assay for detecting quantifying both the direct indirect events result from interactions. The utilizes an H(2) (16)O(2) (18)O(2) strategy to evaluate denaturant dependence of methionine oxidation in proteins presence absence target ligand. differential reactions performed ligand provides measure protein stability changes occur as interactions...