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Threads Started By: nmrlearner
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[NMR paper]
Quantification of Biopharmaceutically Relevant Nonionic Surfactant Excipients Using Benchtop qNMR
Quantification of Biopharmaceutically Relevant Nonionic Surfactant Excipients Using Benchtop qNMR
Nonionic surfactant excipients (NISEs) are commonly added to biologics formulations to mitigate the effects of stress incurred by the active biotherapeutic during manufacturing, transport, and storage. During manufacturing, NISEs are added by dilution of a stock solution directly into a protein formulation, and their accurate addition is critical in maintaining the quality and integrity of the drug product and thus ensuring patient safety. This is especially true for the common NISEs,...
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[NMR paper]
Unraveling the role of probiotics in affecting the structure of monoglyceride gelled emulsions: A low-field (1)H NMR study
Unraveling the role of probiotics in affecting the structure of monoglyceride gelled emulsions: A low-field (1)H NMR study
The capacity of monoglyceride (MG) gelled emulsions (MEs) in protecting probiotic cells of Lacticaseibacillus rhamnosus against stresses suffered during food processing, storage, and human digestion has been recently demonstrated. These findings open new perspectives on the possible participation of probiotics in the stabilization of emulsion structure. To unravel this aspect, rheological analysis and Low-Field ¹H NMR investigations were performed on MEs having...
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[NMR paper]
Dynamics of Coordinated Phosphonate Group Directly Observed by 17O-NMR in Lanthanide(III) Complexes of a Mono(ethyl phosphonate) DOTA Analogue
Dynamics of Coordinated Phosphonate Group Directly Observed by 17O-NMR in Lanthanide(III) Complexes of a Mono(ethyl phosphonate) DOTA Analogue
Biological phosphates can coordinate metal ions and their complexes are common in living systems. Dynamics of mutual oxygen atom exchange in the tetrahedral group in complexes has not been investigated. Here, we present a direct experimental proof of the exchange ("phosphonate rotation") in model Ln(III) complexes of monophosphonate H4dota analogue which alters phosphorus atom chirality of coordinated phosphonate monoester. Combination of...
nmrlearner
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[NMR paper]
DREAMweb: An online tool for graph-based modeling of NMR protein structure
DREAMweb: An online tool for graph-based modeling of NMR protein structure
The value of accurate protein structural models closely conforming to the experimental data is indisputable. DREAMweb deploys an improved DREAM algorithm, DREAMv2, that incorporates a tighter bound in the constraint set of the underlying optimization approach. This reduces the artifacts while modeling the protein structure by solving the distance-geometry problem. DREAMv2 follows a bottom-up strategy of building smaller substructures for regions with a larger concentration of experimental bounds...
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[NMR paper]
SCREENES: Enhancing non-uniform sampling reconstruction for symmetrical NMR spectroscopy
SCREENES: Enhancing non-uniform sampling reconstruction for symmetrical NMR spectroscopy
BACKGROUND: Symmetrical NMR spectroscopy, such as Total Correlation Spectroscopy (TOCSY) and other homonuclear spectroscopy, displays symmetry in chemical shift but are generally not symmetrical in terms of intensity, which constitutes a pivotal branch of multidimensional NMR spectroscopy and offers a robust tool for elucidating the structures and dynamics of complex samples, particularly in the context of biological macromolecules. Non-Uniform Sampling (NUS) stands as a critical technique for...
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[NMR paper]
Structural analysis of ATP bound to the F(1)-ATPase beta-subunit monomer by solid-state NMR- insight into the hydrolysis mechanism in F(1)
Structural analysis of ATP bound to the F(1)-ATPase beta-subunit monomer by solid-state NMR- insight into the hydrolysis mechanism in F(1)
ATP-hydrolysis-associated conformational change of the ?-subunit during the rotation of F(1)-ATPase (F(1)) has been discussed using cryo-electron microscopy (cryo-EM). Since it is worthwhile to further investigate the conformation of ATP at the catalytic subunit through an alternative approach, the structure of ATP bound to the F(1)?-subunit monomer (?) was analyzed by solid-state NMR. The adenosine conformation of ATP-? was similar to that of ATP...
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[NMR paper]
4,4-Difluoroproline as a Unique (19)F NMR Probe of Proline Conformation
4,4-Difluoroproline as a Unique (19)F NMR Probe of Proline Conformation
Despite the importance of proline conformational equilibria (trans versus cis amide and exo versus endo ring pucker) on protein structure and function, there is a lack of convenient ways to probe proline conformation. 4,4-Difluoroproline (Dfp) was identified to be a sensitive ^(19)F NMR-based probe of proline conformational biases and cis-trans isomerism. Within model compounds and disordered peptides, the diastereotopic fluorines of Dfp exhibit similar chemical shifts (??(FF) = 0-3 ppm) when a...
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nmrlearner
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nmrlearner
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[NMR paper]
(17)O Solid-State NMR Spectroscopy of Lipid Membranes
(17)O Solid-State NMR Spectroscopy of Lipid Membranes
Despite the limitations posed by poor sensitivity, studies have reported the unique advantages of ^(17)O based NMR spectroscopy to study systems existing in liquid, solid, or semisolid states. ^(17)O NMR studies have exploited the remarkable sensitivity of quadrupole coupling and chemical shift anisotropy tensors to the local environment in the characterization of a variety of intra- and intermolecular interactions and motion. Recent studies have considerably expanded the use of ^(17)O NMR to...
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[NMR paper]
1D and 2D NMR for KRAS:Ligand Binding
1D and 2D NMR for KRAS:Ligand Binding
Fragment-based screening by ligand-observed 1D NMR and binding interface mapping by protein-observed 2D NMR are popular methods used in drug discovery. These methods allow researchers to detect compound binding over a wide range of affinities and offer a simultaneous assessment of solubility, purity, and chemical formula accuracy of the target compounds and the ^(15)N-labeled protein when examined by 1D and 2D NMR, respectively. These methods can be applied for screening fragment binding to the...
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[NMR paper]
Profiling Complex RAS-Effector Interactions Using NMR Spectroscopy
Profiling Complex RAS-Effector Interactions Using NMR Spectroscopy
Knowledge of how effectors interact with RAS GTPases is key to understanding how these switch-like proteins function in cells. Effectors bind specifically to GTP-loaded RAS using RAS association (RA) or RAS binding domains (RBDs) that show wide-ranging affinities and thermodynamic characteristics. Both normal development and RAS-induced tumorigenesis depend on multiple distinct effector proteins that are frequently co-expressed and co-localized, suggesting an antagonistic nature to signaling...
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[NMR paper]
Real-Time Monitoring of RAS Activity Using In Vitro and In-Cell NMR Spectroscopy
Real-Time Monitoring of RAS Activity Using In Vitro and In-Cell NMR Spectroscopy
The activation level of RAS can be determined by GTP hydrolysis rate (k(hy)) and GDP-GTP exchange rates (k(ex)). Either impaired GTP hydrolysis or enhanced GDP-GTP exchange causes the aberrant activation of RAS in oncogenic mutants. Therefore, it is important to quantify the k(hy) and k(ex) for understanding the mechanisms of RAS oncogenesis and drug development. Conventional methods have individually measured the k(ex) and k(hy) of RAS. However, within the intracellular environment, GTP...
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[NMR paper]
Protocol to identify the ligand binding site of Mincle using NMR spectroscopy
Protocol to identify the ligand binding site of Mincle using NMR spectroscopy
Mincle (macrophage-inducible C-type lectin, CLEC4E) is a C-type lectin immune-stimulatory receptor that can be targeted for inducing potent adjuvant effects. Mincle can recognize trehalose dimycolate and related glycolipids. Here, we present a protocol to identify the ligand binding mode of Mincle. We describe steps for preparing labeled Mincle ectodomain, data acquisition, and analysis of nuclear magnetic resonance experiments using non-detergent sulfobetaine-195. This protocol can be applied...
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[NMR paper]
Assessment of monoclonal antibody glycosylation: a comparative study using HRMS, NMR, and HILIC-FLD
Assessment of monoclonal antibody glycosylation: a comparative study using HRMS, NMR, and HILIC-FLD
Monoclonal antibodies (mAbs) represent the largest class of therapeutic protein drug products. mAb glycosylation produces a heterogeneous, analytically challenging distribution of glycoforms that typically should be adequately characterized because glycosylation-based product quality attributes (PQAs) can impact product quality, immunogenicity, and efficacy. In this study, two products were compared using a panel of analytical methods. Two high-resolution mass spectrometry (HRMS) workflows...
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