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Concept: Conformation


With a view to development of novel sialidase inhibitors, mimetics of the natural inhibitor Neu5Ac2en have been prepared in which a phosphonate group replaces the sialic acid glycerol side chain. Different hex-4-en derivatives adopt half-chair conformations that place the glycosyl phosphonate in an equatorial position. For the α-L-threo-hex-4-en derivative this conformation is equivalent to that of Neu5Ac2en, and opposite to that seen for alkyl O-glycosides with the same overall stereochemistry.

Concepts: Tertiary structure, Stereochemistry, Calculus, Vector space, Derivative, Conformation, Conformational isomerism, Cyclohexane conformation


Nanobodies are single-domain antibodies found in camelids. These are the smallest naturally occurring binding domains and derive functionality via three hypervariable loops (H1, H2 and H3) which form the binding surface. They are excellent candidates for antibody engineering due to their favorable characteristics like small-size, high solubility, and stability. To rationally engineer antibodies with affinity for a specific target, one can tailor the hypervariable loops to obtain the desired binding surface. As a first step toward such a goal, we consider the design of loops with a desired conformation. In this study, we focus on the H1 loop of the anti-hCG llama nanobody which exhibits a non-canonical conformation. We aim to “tilt” the balance of the H1 loop conformation from a non-canonical conformation to a (humanized) type-1 canonical conformation by correlating the effect of selected mutations to the amino-acid sequence of the H1, H2 and proximal residues on the H1 loop conformation. We use all-atomistic, explicit-solvent, biased molecular dynamic simulations to simulate the wildtype and mutant loops in a pre-folded framework. We thus find mutants with increasing propensity to form a stable type-1 canonical conformation of the H1 loop. Free-energy landscapes reveal the existence of conformational isomers of the canonical conformation which may play a role in binding different antigenic surfaces. We also elucidate the approximate mechanism and kinetics of transitions between such conformational isomers by using a Markovian model. We find that a particular three-point mutant has the largest thermodynamic propensity to form the H1 type-1 canonical structure but also to exhibit transitions between conformational isomers, while a different, more rigid three-point mutant has the largest propensity to be kinetically trapped in such a canonical structure.

Concepts: Immune system, Antibody, Monoclonal antibodies, Tertiary structure, Conformation, Conformational isomerism, Isomer, Single domain antibody


Efficient synthesis of tetraoxacalix[2]perfluoroarene[2]triazines and their isomeric analogs from a one-pot macrocyclic condensation reaction of methoxy- and amino-substituted dichlorotriazines with tetrafluorobenzene-1,3-, -1,4-, and -1,2-diols was developed. X-ray analysis demonstrates that they adopt drastically different 1,3-alternate conformations in the crystalline state while in solution they undergo very fast conformational changes relative to the NMR time scale.

Concepts: Fundamental physics concepts, Chemical synthesis, Conformation


Single-molecule pulling experiments on unstructured proteins linked to neurodegenerative diseases have measured rupture forces comparable to those for stable folded proteins. To investigate the structural mechanisms of this unexpected force resistance, we perform pulling simulations of the amyloid β-peptide (Aβ) and α-synuclein (αS), starting from simulated conformational ensembles for the free monomers. For both proteins, the simulations yield a set of rupture events that agree well with the experimental data. By analyzing the conformations occurring shortly before rupture in each event, we find that the mechanically resistant structures share a common architecture, with similarities to the folds adopted by Aβ and αS in amyloid fibrils. The disease-linked Arctic mutation of Aβ is found to increase the occurrence of highly force-resistant structures. Our study suggests that the high rupture forces observed in Aβ and αS pulling experiments are caused by structures that might have a key role in amyloid formation.

Concepts: Causality, Structure, Tertiary structure, Prion, Neurodegeneration, Engineering, Simulation, Conformation


High temperature is known to cause some instability in polysaccharide-protein conjugated vaccines and studies under stress conditions may be useful in determining whether short-term accidental exposure to undesired conditions can compromise product quality. In this study, we examined the structural stability of three industrial batches of Brazilian Meningococcal C conjugate bulk (MPCT) incubated at 4, 37, and 55 °C for 5 weeks. The effect of exposure to the storage temperatures was monitored by HPLC-SEC, CZE, CD and NMR techniques. The immunological significance of any physicochemical changes observed in MPCT was determined by SBA and ELISA assays of serum from immunized mice. Fluorescence emission spectra at 4 and 37 °C were similar among all samples and compatible with the native fold of the carrier protein. Fluorescence spectra of MPCT stored at 55 °C decreased in intensity and had a significant red-shift, indicating conformational changes. Far-UV CD spectra revealed a trend toward loss of structural conformation as storage temperature was increased to 55 °C. The NMR data showed modified signal intensity of the aromatic and aliphatic residues, mainly for samples incubated at 55 °C, suggesting a partial loss of tertiary structure. About 50% free saccharide content was found in bulks stored at 55 °C, but no difference was observed in the IgG or SBA titers. The present study showed physicochemical methods alone are insufficient to predict the biological activity of a MPCT conjugate vaccine without extensive validation against immunological data. However, they provide a sensitive means of detecting changes induced in a vaccine exposed to adverse environmental condition.

Concepts: Immune system, Fluorescence, Vaccination, Light, Immunology, Temperature, ELISA, Conformation


ATP-phosphoribosyltransferase (ATP-PRT) is a hexameric enzyme in conformational equilibrium between an open and seemingly active state and a closed and presumably inhibited form. The structure-function relationship of allosteric regulation in this system is still not fully understood. Here, we develop a screening strategy for modulators of ATP-PRT and identify 3-(2-thienyl)-L-alanine (TIH) as an allosteric activator of this enzyme. Kinetic analysis reveals co-occupancy of the allosteric sites by TIH and L-histidine. Crystallographic and native ion-mobility mass spectrometry data show that the TIH-bound activated form of the enzyme closely resembles the inhibited L-histidine-bound closed conformation, revealing the uncoupling between ATP-PRT open and closed conformations and its functional state. These findings suggest that dynamic processes are responsible for ATP-PRT allosteric regulation and that similar mechanisms might also be found in other enzymes bearing a ferredoxin-like allosteric domain.Active and inactive state ATP-phosphoribosyltransferases (ATP-PRTs) are believed to have different conformations. Here the authors show that in both states, ATP-PRT has a similar structural arrangement, suggesting that dynamic alterations are involved in ATP-PRT regulation by allosteric modulators.

Concepts: Allosteric regulation, Proteins, Enzyme, Tertiary structure, Chemical equilibrium, U.S. state, State, Conformation


The question of how many chains an elementary cellulose microfibril contains is critical to understanding the molecular mechanism(s) of cellulose biosynthesis and regulation. Given the hexagonal nature of the cellulose synthase rosette it is assumed that the number of chains must be a multiple of six. We present molecular dynamics simulations on three different models of Iβ cellulose microfibrils, 18, 24 and 36 chains, to investigate their structure and dynamics in a hydrated environment. The 36 chain model stays in a conformational space that is very similar to the initial crystalline phase while the 18 and 24 chain models sample a conformational space different to the crystalline structure, yet similar to conformations observed in recent high temperature MD simulations. Major differences in the conformations sampled between the different models result from changes to the tilt of chains in different layers, specifically a second stage of tilt; increased rotation about the O2-C2 dihedral; and a greater sampling of non-TG exocyclic conformations, particularly the GG conformation in center layers and GT conformation in solvent exposed exocyclic groups. With a re-interpretation of NMR data, specifically for contributions made to the C6 peak, data from the simulations suggest that the 18 and 24 chain structures are more viable models for an elementary cellulose microfibril, which also correlates with recent scattering and diffraction experimental data. These data inform biochemical and molecular studies that must explain how a six particle cellulose synthase complex rosette synthesises microfibrils likely comprised of either 18 or 24 chains.

Concepts: Molecular dynamics, Tertiary structure, Chemistry, Atom, Computational chemistry, Difference, Cellulose, Conformation


Ensemble docking corresponds to the generation of an “ensemble” of drug target conformations in computational structure-based drug discovery, often obtained by using molecular dynamics simulation, that is used in docking candidate ligands. This approach is now well established in the field of early-stage drug discovery. This review gives a historical account of the development of ensemble docking and discusses some pertinent methodological advances in conformational sampling.

Concepts: Bioinformatics, Molecular dynamics, Computational chemistry, Drug design, Monte Carlo method, Computer simulation, Conformation, Molecular mechanics


Concern has arisen in recent years that selection for extreme facial morphology in the domestic dog may be leading to an increased frequency of eye disorders. Corneal ulcers are a common and painful eye problem in domestic dogs that can lead to scarring and/or perforation of the cornea, potentially causing blindness. Exaggerated juvenile-like craniofacial conformations and wide eyes have been suspected as risk factors for corneal ulceration. This study aimed to quantify the relationship between corneal ulceration risk and conformational factors including relative eyelid aperture width, brachycephalic (short-muzzled) skull shape, the presence of a nasal fold (wrinkle), and exposed eye-white. A 14 month cross-sectional study of dogs entering a large UK based small animal referral hospital for both corneal ulcers and unrelated disorders was carried out. Dogs were classed as affected if they were diagnosed with a corneal ulcer using fluorescein dye while at the hospital (whether referred for this disorder or not), or if a previous diagnosis of corneal ulcer(s) was documented in the dogs' histories. Of 700 dogs recruited, measured and clinically examined, 31 were affected by corneal ulcers. Most cases were male (71%), small breed dogs (mean± SE weight: 11.4±1.1 kg), with the most commonly diagnosed breed being the Pug. Dogs with nasal folds were nearly five times more likely to be affected by corneal ulcers than those without, and brachycephalic dogs (craniofacial ratio <0.5) were twenty times more likely to be affected than non-brachycephalic dogs. A 10% increase in relative eyelid aperture width more than tripled the ulcer risk. Exposed eye-white was associated with a nearly three times increased risk. The results demonstrate that artificially selecting for these facial characteristics greatly heightens the risk of corneal ulcers, and such selection should thus be discouraged to improve canine welfare.

Concepts: Eye, Fluorescein, Contact lens, Dog, Dog breed, Corneal ulcer, Conformation, Gray Wolf


Intrinsically disordered proteins dynamically sample a wide conformational space and therefore do not adopt a stable and defined three-dimensional conformation. The structural heterogeneity is related to their proper functioning in physiological processes. Knowledge of the conformational ensemble is crucial for a complete comprehension of this kind of proteins. We here present an approach that utilizes dynamic nuclear polarization-enhanced solid-state NMR spectroscopy of sparsely isotope-labeled proteins in frozen solution to take snapshots of the complete structural ensembles by exploiting the inhomogeneously broadened line-shapes. We investigated the intrinsically disordered protein α-synuclein (α-syn), which plays a key role in the etiology of Parkinson’s disease, in three different physiologically relevant states. For the free monomer in frozen solution we could see that the so-called “random coil conformation” consists of α-helical and β-sheet-like conformations, and that secondary chemical shifts of neighboring amino acids tend to be correlated, indicative of frequent formation of secondary structure elements. Based on these results, we could estimate the number of disordered regions in fibrillar α-syn as well as in α-syn bound to membranes in different protein-to-lipid ratios. Our approach thus provides quantitative information on the propensity to sample transient secondary structures in different functional states. Molecular dynamics simulations rationalize the results.

Concepts: Protein, Protein structure, Amino acid, Secondary structure, Structure, Tertiary structure, Polymer, Conformation