A5055110638- Mass Spectrometry Techniques and Applications
- Analytical Chemistry and Chromatography
- Advanced Proteomics Techniques and Applications
- Protein purification and stability
- Monoclonal and Polyclonal Antibodies Research
- Protein Structure and Dynamics
- Enzyme Structure and Function
- Biochemical and Molecular Research
- Ion-surface interactions and analysis
- Advanced ceramic materials synthesis
- Analytical Chemistry and Sensors
- RNA and protein synthesis mechanisms
- Pharmacogenetics and Drug Metabolism
- HER2/EGFR in Cancer Research
- Metabolomics and Mass Spectrometry Studies
- Glycosylation and Glycoproteins Research
- Receptor Mechanisms and Signaling
- Molecular Junctions and Nanostructures
- Pancreatic function and diabetes
- Neonatal Health and Biochemistry
- Tuberculosis Research and Epidemiology
- Heme Oxygenase-1 and Carbon Monoxide
- Viral Infectious Diseases and Gene Expression in Insects
- Calcium signaling and nucleotide metabolism
- Methemoglobinemia and Tumor Lysis Syndrome
Pharmaron (United Kingdom)
2025
University of Manchester
2015-2024
Protéomique, Réponse Inflammatoire et Spectrométrie de Masse
2019
University of Edinburgh
2012-2017
In the past decade, mass spectrometry (MS) coupled with electrospray ionization (ESI) has been extensively applied to study of intact proteins and their complexes, often without requirement labels. Solvent conditions (for example, pH, ionic strength, concentration) affect observed desolvated species; ease altering such extrinsic factors renders ESI-MS an appropriate method by which consider range conformational states that may occupy, including natively folded, disordered amyloid....
Abstract Collision cross‐sections (CCS) of immunoglobulins G1 and G4 have been determined using linear drift‐tube ion‐mobility mass spectrometry. Intact antibodies Fc‐hinge fragments present with a larger range CCS than proteins comparable size. This is rationalized MD simulations, which indicate significant in vacuo dynamics between linked folded domains. The IgG4 subclass presents over wider the IgG1 subclass.
Antibody-drug-conjugates (ADC) are a growing class of anticancer biopharmaceuticals. Conjugation cysteine linked ADCs, requires initial reduction mAb inter-chain disulfide bonds, as the drugs attached via thiol chemistry. This results in active moiety being transformed from covalently tetramer to non-covalently complexes, which hinders precise determination drug load with LC-MS. Here, we show how addition charge reducing agent triethylammonium acetate (TEAA) preserves intact structure, is...
Progressive decline in functional beta cell mass is central to the development of type 2 diabetes. Elevated serum levels extracellular nicotinamide phosphoribosyltransferase (eNAMPT) are associated with failure diabetes and eNAMPT immuno-neutralisation improves glucose tolerance mouse models Despite this, effects on poorly elucidated, some studies having separately reported cell-protective eNAMPT. exists structurally functionally distinct monomeric dimeric forms. Dimerisation essential for...
Adeno-associated viruses (AAVs) are at the forefront of biopharmaceutical development as gene therapy vectors. The successful approval these medicines requires robust characterization including an assessment therapeutic content. Using ion mobility–mass spectrometry (IM-MS) techniques, we determine empty:full capsid ratios and explore structural differences in AAV particles relevant to payload. With drift tube IM-MS, demonstrate that empty capsids, while slightly smaller, present more...
The thermal stability and strength of interactions in proteins are commonly measured using isothermal calorimetry differential scanning providing a measurement that averages over structural transitions occur as the melt dissociate. Here, we apply variable temperature ion mobility mass spectrometry (VT-IM-MS) to study effect on structure four multimeric protein complexes. VT-IM-MS is used here investigate change conformation model proteins, namely, transthyretin (TTR), avidin, concanavalin A...
Thermally induced conformational transitions of three proteins increasing intrinsic disorder-cytochrome c, the tumor suppressor protein p53 DNA binding domain (p53 DBD), and N-terminus oncoprotein murine double minute 2 (NT-MDM2)-have been studied by native mass spectrometry variable-temperature drift time ion mobility (VT-DT-IM-MS). Ion measurements were carried out at temperatures ranging from 200 to 571 K. Multiple conformations are observable over several charge states for all monomeric...
To quantify the measurable structural heterogeneity of a biopharmaceutical product and effect glycosylation on this we systematically evaluate three lots Herceptin®, two mAb standards an intact 5 Fc-hinge fragment.
The aggregation of protein-based therapeutics such as monoclonal antibodies (mAbs) can affect the efficacy treatment and even induce effects that are adverse to patient. Protein engineering is used shift mAb away from an aggregation-prone state by increasing thermodynamic stability native fold, which might in turn alter conformational flexibility. We have probed thermal three types intact IgG molecules two Fc-hinge fragments using variable-temperature ion-mobility mass spectrometry...
Charge reduction in the gas phase provides a direct means of manipulating protein charge state, and when coupled to ion mobility mass spectrometry (IM-MS), it is possible monitor effect on conformation absence solution. Use electron transfer reagent 1,3-dicyanobenzene, with IM-MS, allows us two proteins deliberately chosen from opposite sides order disorder continuum: bovine pancreatic trypsin inhibitor (BPTI) beta casein. The ordered BPTI presents compact conformers for each three states...
Abstract ATP-phosphoribosyltransferase (ATP-PRT) is a hexameric enzyme in conformational equilibrium between an open and seemingly active state closed presumably inhibited form. The structure-function relationship of allosteric regulation this system still not fully understood. Here, we develop screening strategy for modulators ATP-PRT identify 3-(2-thienyl)- l- alanine (TIH) as activator enzyme. Kinetic analysis reveals co-occupancy the sites by TIH l -histidine. Crystallographic native...
Abstract Collision cross‐sections (CCS) of immunoglobulins G1 and G4 have been determined using linear drift‐tube ion‐mobility mass spectrometry. Intact antibodies Fc‐hinge fragments present with a larger range CCS than proteins comparable size. This is rationalized MD simulations, which indicate significant in vacuo dynamics between linked folded domains. The IgG4 subclass presents over wider the IgG1 subclass.
The Mycobacterium tuberculosis (Mtb) heme oxygenase MhuD liberates free iron by degrading to the linear tetrapyrrole mycobilin. dimer binds up two hemes within active site of each monomer. Binding first solvent-exposed allows degradation and releases iron. a second renders inactive, allowing storage. Native-mass spectrometry revealed little difference in binding affinity between solvent-protected hemes. Hence, diheme-MhuD is formed even when large proportion population apo form. Apomyoglobin...
The cytochrome P450 monooxygenase BM3 (BM3) is a biotechnologically important and versatile enzyme capable of producing compounds such as the medical drugs pravastatin artemether, steroid hormone testosterone. natural fusion comprising two major domains: (heme-binding) catalytic domain NADPH-cytochrome reductase (CPR) containing FAD FMN cofactors in distinct domains CPR. A crystal structure full-length not available its monomeric or catalytically active dimeric state. In this study, we...
Measurements of protein higher order structure (HOS) provide important information on stability, potency, efficacy, immunogenicity, and biosimilarity biopharmaceuticals, with a significant number techniques methods available to perform these measurements. The comparison the analytical performance HOS standardization results is, however, not trivial task, due lack reference protocols measurement procedures. Here, we developed protocol structurally alter compare samples somatropin, recombinant...
<title>Abstract</title> The increasing prevalence of antimicrobial resistance is an important challenge that warrants new approaches to antibiotic development. Currently, all antibiotics inhibit biological processes. To explore whether activation a biochemical pathway can elicit bactericidal effects we engineered variants Mycobacterium tuberculosis ATP-phosphoribosyltransferase (ATP-PRT) are resistant allosteric inhibition by L-histidine, leading supraphysiological ATP-PRT and L-histidine...
Abstract Review: 204 refs.
<p>To consider the measurable variations in biopharmaceuticals we use mass spectrometry and systematically evaluate three lots of Herceptin®, two mAb standards an intact Fc-hinge fragment. Each is examined states; glycan intact, truncated (following endoS2 treatment) fully deglycosylated. Despite equivalence at protein level, each lot Herceptin® gives a distinctive signature different analyses. Ion mobility (IM-MS) shows that API, attached N-glycans reduce conformational spread by 10.5...
To consider the measurable variations in biopharmaceuticals we use mass spectrometry and systematically evaluate three lots of Herceptin®, two mAb standards an intact Fc-hinge fragment. Each is examined states; glycan intact, truncated (following endoS2 treatment) fully deglycosylated. Despite equivalence at protein level, each lot Herceptin® gives a distinctive signature different analyses. Ion mobility (IM-MS) shows that API, attached N-glycans reduce conformational spread by 10.5 – 25 %....