Backbone Conformation Research Articles

HLA3DB: comprehensive annotation of peptide/HLA complexes enables blind structure prediction of T cell epitopes

The class I proteins of the major histocompatibility complex (MHC-I) display epitopic peptides derived from endogenous proteins on the cell surface for immune surveillance. Accurate modeling of peptides bound to the human MHC, HLA, has been mired by conformational diversity of the central peptide residues, which are critical for recognition by T cell receptors. Here, analysis of X-ray crystal structures within our curated database (HLA3DB) shows that pHLA complexes encompassing multiple HLA allotypes present a discrete set of peptide backbone conformations. Leveraging these backbones, we employ a regression model trained on terms of a physically relevant energy function to develop a comparative modeling approach for nonamer pHLA structures named RepPred. Our method outperforms the top pHLA modeling approach by up to 19% in structural accuracy, and consistently predicts blind targets not included in our training set. Insights from our work may be applied towards predicting antigen immunogenicity, and receptor cross-reactivity.

The pseudotorsional space of RNA.

The characterization of the conformational landscape of the RNA backbone is rather complex due to the ability of RNA to assume a large variety of conformations. These backbone conformations can be depicted by pseudotorsional angles linking RNA backbone atoms, from which Ramachandran-like plots can be built. We explore here different definitions of these pseudotorsional angles, finding that the most accurate ones are the traditional η (eta) and θ (theta) angles, which represent the relative position of RNA backbone atoms P and C4'. We explore the distribution of η - θ in known experimental structures, comparing the pseudotorsional space generated with structures determined exclusively by one experimental technique. We found that the complete picture only appears when combining data from different sources. The maps provide a quite comprehensive representation of the RNA accessible space, which can be used in RNA-structural predictions. Finally, our results highlight that protein interactions lead to significant changes in the population of the η - θ space, pointing toward the role of induced-fit mechanisms in protein-RNA recognition.

Benzotriazole‐Based D–π–A‐Type Photovoltaic Polymers Break Through 17% Efficiency

AbstractBenzo[d][1,2,3]triazole (BTA) unit is one of the most classic electron‐accepting units (A) to construct donor (D)‐π‐A‐type photovoltaic polymers. However, the highest power conversion efficiency (PCE) of organic photovoltaics (OPVs) based on BTA‐containing polymers is restricted to 15–16%, lagging other promising polymers. Thus, investigating the structure‐performance relationship and breaking the efficiency bottleneck of BTA‐based polymers is challenging but critical. Herein, the effects of fusing two thiophene rings at D (PE52), π (PE4), and A (PE39) units of a classic D–π–A‐type BTA‐containing polymer J52‐Cl, respectively, on the backbone conformation, crystallinity, molecular stacking, and photovoltaic performance are systematically investigated. When blended with a BTA‐containing non‐fullerene acceptor (NFA), Y18, all three polymers with extending conjugated backbones can decrease the energy loss of photovoltaic devices. Notably, PE4, with a linear backbone conformation, realizes the champion PCE of 17.08%, with a short‐circuit current density (JSC) of 26.83 mA cm−2, a large breakthrough for BTA‐based photovoltaic polymers. What's more, the photovoltaic device based on PE4:Y18 combination fabricated by a non‐halogenated solvent of o‐xylene also displays an excellent PCE of 16.87%. The results indicate that fusing thiophene rings to BTA‐polymers, especially at π‐bridge, is a simple and effective method to improve the photovoltaic performance via modulating the molecular conformation and crystallinity.

Interfacial shear rheology of glassy polymers at liquid interfaces

When surface-active molecules or particles assemble at fluid–fluid interfaces, these interfaces acquire complex rheological properties that are of importance in processes that involve flow and deformation of interfaces. Although much progress has been made, interfacial rheology measurements and, in particular, the measurement of interfacial rheological properties of polymers at the air-water interface remain challenging. These are due to weak interactions with the water subphase, the polymer backbone conformation, the glass transition of the interfacial layer, and memory effects. In the present work, we describe systematic rheological measurements of polymer-laden interfaces. The measurements were performed with four different interfacial shear rheometers that can be classified into two types: rheometers in which the surface pressure can be controlled independently, and devices based on fixtures mounted on standard rotational rheometers and lacking control of the surface pressure. We use poly(tert-butyl methacrylate) and poly(methyl methacrylate), two high glass transition temperature, hydrophobic polymers anchored to the water subphase by means of the acrylate group. Using a Langmuir–Pockels (LP) trough, we identify the transition of the polymer monolayer from a viscous to a solid elastic or soft-glassy interface as the polymer surface concentration increases by compression. Then, we compare the linear viscoelastic properties of the interface as obtained by each rheometer. Our results show poor reproducibility and comparability of the rheological data as obtained by different rheometers for the same polymer. This is mainly due to differences in the method used to prepare the layers. For LP-based devices, spreading under dilute conditions and subsequent compression yields layers of compressed glassy blobs with reproducible results. On the other hand, for devices without surface pressure control, deposition of the amount needed to reach a desired concentration may lead to the formation of ill-defined layers resulting in irreproducible data. Furthermore, we find that only when spreading the polymer to form a dilute layer and then controlling the surface pressure by compression, we can clearly distinguish the fluidlike from solidlike interfaces, and a clear correlation is observed between the surface pressure (or interfacial polymer concentration) and the rheological properties of the interface.

Discovery and rational engineering of PET hydrolase with both mesophilic and thermophilic PET hydrolase properties

Excessive polyethylene terephthalate (PET) waste causes a variety of problems. Extensive research focused on the development of superior PET hydrolases for PET biorecycling has been conducted. However, template enzymes employed in enzyme engineering mainly focused on IsPETase and leaf-branch compost cutinase, which exhibit mesophilic and thermophilic hydrolytic properties, respectively. Herein, we report a PET hydrolase from Cryptosporangium aurantiacum (CaPETase) that exhibits high thermostability and remarkable PET degradation activity at ambient temperatures. We uncover the crystal structure of CaPETase, which displays a distinct backbone conformation at the active site and residues forming the substrate binding cleft, compared with other PET hydrolases. We further develop a CaPETaseM9 variant that exhibits robust thermostability with a Tm of 83.2 °C and 41.7-fold enhanced PET hydrolytic activity at 60 °C compared with CaPETaseWT. CaPETaseM9 almost completely decompose both transparent and colored post-consumer PET powder at 55 °C within half a day in a pH-stat bioreactor.

Exploring binding positions and backbone conformations of peptide ligands of proteins with a backbone-centred statistical energy function.

When designing peptide ligands based on the structure of a protein receptor, it can be very useful to narrow down the possible binding positions and bound conformations of the ligand without the need to choose its amino acid sequence in advance. Here, we construct and benchmark a tool for this purpose based on a recently reported statistical energy model named SCUBA (Sidechain-Unknown Backbone Arrangement) for designing protein backbones without considering specific amino acid sequences. With this tool, backbone fragments of different local conformation types are generated and optimized with SCUBA-driven stochastic simulations and simulated annealing, and then ranked and clustered to obtain representative backbone fragment poses of strong SCUBA interaction energies with the receptor. We computationally benchmarked the tool on 111 known protein-peptide complex structures. When the bound ligands are in the strand conformation, the method is able to generate backbone fragments of both low SCUBA energies and low root mean square deviations from experimental structures of peptide ligands. When the bound ligands are helices or coils, low-energy backbone fragments with binding poses similar to experimental structures have been generated for approximately 50% of benchmark cases. We have examined a number of predicted ligand-receptor complexes by atomistic molecular dynamics simulations, in which the peptide ligands have been found to stay at the predicted binding sites and to maintain their local conformations. These results suggest that promising backbone structures of peptides bound to protein receptors can be designed by identifying outstanding minima on the SCUBA-modeled backbone energy landscape.

Stable Deep‐Blue Electroluminescence from Hierarchical Crystalline Structure Based on Planar Conformational Terfluorenes: Benefits of Preventing Defect Structures Formation

AbstractLiner‐type organic conjugated molecules with 1D π‐electron delocalization present conformation‐dependent intramolecular optoelectronic behavior. Significantly different from non‐planar ones, the coplanar conformational backbone of π‐conjugated molecules can enhance intramolecular π‐electron delocalization and dominate intermolecular π‐electron coupling, which are the fundamental parameters for improving their optoelectronic properties. Herein, the effect of the formation of coplanar conformation on intermolecular packing and optoelectrical properties of liner‐type steric terfluorene (DSFX‐SFSO) toward solution‐processed deep‐blue organic light‐emitting diodes (OLEDs) is systematically demonstrated. Firstly, uniform and continuous crystalline films based on the planar conformational DSFX‐SFSO molecules are achieved via thermal annealing at 160 °C, which can avoid microscale crystallization processing. Contrary to amorphous ones, these hierarchical crystalline films present robust deep‐blue emission with an efficiency of > 60%, due to the suppression of defect structures. More interestingly, this hierarchical crystalline film has a stable and efficient deep‐blue electroluminescence (CIE: 0.15, 0.09) without the obvious defective green‐band emission, even at 500 mA cm−2, also further confirming the excellent and perfect molecular landscape to obtain single‐chromophore excitonic behavior from single planar‐terfluorene segments. Therefore, obtaining a uniform hierarchical structure based on planar conformational molecules is an effective strategy to optimize the optoelectrical properties for optoelectronic applications.

Monitoring Conformation and Protonation States of Glutathione by Raman Optical Activity and Molecular Dynamics.

Glutathione (GSH) is a common antioxidant and its biological activity depends on the conformation and protonation state. We used molecular dynamics, Raman and Raman optical activity (ROA) spectroscopies to investigate GSH structural changes in a broad pH range. Factor analysis of the spectra provided protonation constants (2.05, 3.45, 8.62, 9.41) in good agreement with previously published values. Following the analysis, spectra of differently protonated forms were obtained by extrapolation. The complete deprotonation of the thiol group above pH 11 was clearly visible in the spectra; however, many spectral features did not change much with pH. Experimental spectra at various pH values were decomposed into the simulated ones, which allowed us to study the conformer populations and quality of molecular dynamics (MD). According to this combined ROA/MD analysis conformation of the GSH backbone is affected by the pH changes only in a limited way. The combination of ROA with the computations thus has the potential to improve the MD force field and obtain more accurate populations of the conformer species. The methodology can be used for any molecule, but for a more detailed insight better computational techniques are needed in the future.

Energy Landscapes and Heat Capacity Signatures for Monomers and Dimers of Amyloid-Forming Hexapeptides.

Amyloid formation is a hallmark of various neurodegenerative disorders. In this contribution, energy landscapes are explored for various hexapeptides that are known to form amyloids. Heat capacity (CV) analysis at low temperature for these hexapeptides reveals that the low energy structures contributing to the first heat capacity feature above a threshold temperature exhibit a variety of backbone conformations for amyloid-forming monomers. The corresponding control sequences do not exhibit such structural polymorphism, as diagnosed via end-to-end distance and a dihedral angle defined for the monomer. A similar heat capacity analysis for dimer conformations obtained using basin-hopping global optimisation shows clear features in end-to-end distance versus dihedral correlation plots, where amyloid-forming sequences exhibit a preference for larger end-to-end distances and larger positive dihedrals. These results hold true for sequences taken from tau, amylin, insulin A chain, a de novo designed peptide, and various control sequences. While there is a little overall correlation between the aggregation propensity and the temperature at which the low-temperature CV feature occurs, further analysis suggests that the amyloid-forming sequences exhibit the key CV feature at a lower temperature compared to control sequences derived from the same protein.

Colorimetric Transition of Polydiacetylene/Cyclodextrin Supramolecular Assemblies and Implications as Colorimetric Sensors for Food Phenolic Antioxidants

AbstractMolecular self‐assembly has significant potential in the field of sensing. Polydiacetylenes (PDAs) are conjugated polymers possessing peculiar optical properties obtained by photopolymerization of self‐assembled diacetylene monomers. Herein, the blue‐to‐red phase transition upon either thermal stimulus or interaction with cyclodextrins (CDs) of two PDAs, bearing either carboxylic (PCDA) or amino (PCDA‐NH2) polar heads, is investigated to develop a colorimetric sensor for food phenolic antioxidants. The change in the PDA polar head does not affect significantly thermo‐chromatic transition. Upon thermal stimulus, in both PDAs, color transition occurs straightforward between two distinct stable states and does not involve the disordering of the PDA crystal phase, as revealed by UV–vis spectroscopy and SAXS analysis. Contrarily, PDA/α‐CD interaction is influenced by intermolecular forces among PDA polar heads and is more efficient for PCDA. α‐CDs presumably cause changes in both PDA backbone conformation and local environment surrounding the individual PDA chains. The PCDA/α‐CD assemblies are investigated as colorimetric sensors for the detection of Tyrosol (Ty) and caffeic acid (CAF), by using the principle of competitive inclusion complex formation. The system results to be more sensitive to CAF than Ty and may permit the determination of CAF in concentration ranges suitable for different food products.

Teraryl Braces in Macrocycles: Synthesis and Conformational Landscape Remodeling of Peptides.

The three-dimensional structure of medium-sized cyclic peptides accounts for their biological activity and other important physiochemical properties. Despite significant advances in the past few decades, chemists' ability to fine-tune the structure, in particular, the backbone conformation, of short peptides made of canonical amino acids is still quite limited. Nature has shown that cross-linking the aromatic side chains of linear peptide precursors via enzyme catalysis can generate cyclophane-braced products with unusual structures and diverse activities. However, the biosynthetic path to these natural products is challenging to replicate in the synthetic laboratory using practical chemical modifications of peptides. Herein, we report a broadly applicable strategy to remodel the structure of homodetic peptides by cross-linking the aromatic side chains of Trp, His, and Tyr residues with various aryl linkers. The aryl linkers can be easily installed via copper-catalyzed double heteroatom-arylation reactions of peptides with aryl diiodides. These aromatic side chains and aryl linkers can be combined to form a large variety of assemblies of heteroatom-linked multi-aryl units. The assemblies can serve as tension-bearable multijoint braces to modulate the backbone conformation of peptides as an entry to previously inaccessible conformational space.

Bistable Resistive Memory Switches fabricated by Floating Thin Films of Conjugated Polymers

The layer-by-layer (LBL) fabrication of conjugated polymer (CPs) thin films while preserving their microstructural features by solution processing is highly desired for compact and flexible electronic circuits. However, it is cumbersome and challenging owing to the unavoidable damage to the underlying layers. To circumvent this issue, the unidirectional floating-film transfer method (UFTM) was utilized for the LBL fabrication of oriented CP thin films on the orthogonal liquid surfaces. Further, resistive bistable memory switches were fabricated by sandwiching a layer of metal nanostructures between the LBL-fabricated oriented CP thin films. The resistive switching phenomena were realized by utilizing the applied bias-dependent charge trapping, holding, and recombination on the available states at vacuum-deposited aluminum nanostructures. The effect of CP backbone conformation on the vertical charge transport was also analyzed via a comparative study of three thiophene-based polymers namely RR-P3HT, PBTTT, and PTB-7. It was revealed that CPs with relatively fewer hydrophobic side chains was more favorable for the facile vertical charge transport due to its preferred face-on conformation on the hydrophilic liquid substrate used in UFTM. It was demonstrated that a non-volatile bistable resistive memory switch fabricated using UFTM-processed oriented thin films of PTB-7 exhibited a remarkably high on-off ratio of 1.5 × 106 with high durability.

The Y46D Mutation Destabilizes Dense Packing of the Second Greek Key Pair of Human γC-Crystallin Causing Congenital Nuclear Cataracts.

The γ-crystallins are highly expressed structural lens proteins comprising four Greek key motifs arranged in two domains. Their globular structure and short-range spatial ordering are essential for lens transparency. Aromatic residues play a vital role in stabilizing Greek key folds by forming Greek key or non-Greek key pairs or tyrosine corners. We investigated the effects of the cataractogenic Y46D mutation in the second Greek key pair (Y46-Y51) of human γC-crystallin on its stability and aggregation. Wild-type and Y46D mutant human γC-crystallin were overexpressed in E. coli BL-21(DE3) PLysS cells, purified using ion-exchange and size-exclusion chromatography, and analyzed by fluorescence spectroscopy and circular dichroism spectroscopy. The Y46D mutation does not affect the γC-crystallin backbone conformation under benign conditions but alters the tryptophan microenvironment, exposing hydrophobic residues to the surface. The Y46D mutant undergoes a three-state transition under thermal stress with midpoints of 54.6 and 67.7 °C while the wild type shows a two-state transition with a midpoint of 77.6 °C. The Y46D mutant also shows a three-state transition under GuHCl stress with Cm values of 0.9 and 2.1 M while the wild type shows a two-state transition with a Cm of 2.4 M GuHCl. Mutant but not wild-type γC-crystallin forms light scattering particles upon heating at 65 °C. Overall, the Y46D CRYGS mutation leaves the protein fold intact under benign conditions but destabilizes the molecule by altering the tryptophan microenvironment and exposing hydrophobic residues to its surface, thus increasing its susceptibility to thermal and chemical stress with resultant self-aggregation, light scattering, and cataract.

Orthogonal Supramolecular Assemblies Using Side-Chain Functionalized Helical Poly(isocyanide)s.

Mimicking the structure of proteins using synthetic polymers requires building blocks with structural similarity and the use of various noncovalent and dynamic covalent interactions. We report the synthesis of helical poly(isocyanide)s bearing diaminopyridine and pyridine side-chains and the multistep functionalization of the polymers' side-chains using hydrogen bonding and metal coordination. The orthogonality of the hydrogen bonding and metal coordination was proved by varying the sequence of the multistep assembly. The two side-chain functionalizations are reversible through the use of competitive solvents and/or competing ligands. Throughout the assembly and disassembly, the helical conformation of the polymer backbone is sustained as proved by circular dichroism spectroscopy. These results open the possibility to incorporate helical domains into complex polymer architectures and create a helical scaffold for smart materials.

Synthesis and Structural Investigation of Foldamer-Based Iodine-Supported Artificial Glycosidases.

Glycosidases are a type of enzyme that hydrolytically cleave carbohydrates and form glycans for biologically important processes. The inadequacies of glycosidases or their genetic abnormalities are responsible for various diseases. Thus, the development of glycosidase mimetics is of great importance. We have designed and synthesized an enzyme mimetic containing l-phenylalanine, α-aminoisobutyric acid (Aib), l-leucine, and m-Nifedipine. From X-ray crystallography, the foldamer adopts a β-hairpin conformation stabilized by two 10-member and one 18-member NH⋅⋅⋅O=C hydrogen bonds. Moreover, the foldamer was found to be highly efficient in hydrolysing ethers and glycosides in the presence of iodine at room temperature. Further, X-ray analysis shows the backbone conformation of the enzyme mimetic to be almost unchanged after the glycosidase reaction. This is the first example of iodine-supported artificial glycosidase activity with an enzyme mimic at ambient conditions.

Decoupling peptide binding from T cell receptor recognition with engineered chimeric MHC-I molecules

Abstract Human leucocyte antigens (HLAs) display a repertoire of epitopic peptides on the cell surface for T cell recognition. Amino acid polymorphisms in the peptide-binding groove of different HLA allotypes result in distinct antigenic repertoires, providing a basis for adaptive immunity to emerging pathogens at the population level. T cell receptors (TCRs) recognize pHLA antigens through interactions with both peptide and heavy chain residues. In this work, we use structural data from peptide:MHC-I and pMHC:TCR structures to identify residues important for peptide and/or TCR binding, and we outline a fixed-backbone computational design approach for engineering synthetic molecules that combine peptide binding and TCR recognition surfaces from existing HLA allotypes. X-ray crystallography demonstrates that chimeric molecules bridging divergent HLA alleles can bind selected peptide antigens in a specified backbone conformation. Moreover, in vitro tetramer staining and biophysical binding experiments using chimeric pMHC-I molecules presenting established antigens further demonstrate the requirement of TCR recognition on interactions with HLA framework residues, as opposed to interactions with peptide-centric Chimeric Antigen Receptors (CARs). Our results underscore a novel, structure-guided platform for developing synthetic HLA molecules displaying tumor-associated antigens with modified TCR interaction surfaces, which could further serve as platforms to elicit alloreactive T cell responses against self-antigens that are overexpressed in cancer. Finally, we gain a deeper understanding of the evolution and functional divergence of the different HLA allotypes by demonstrating an example of convergent evolution. Supported by grants from NIAID (5R01AI143997), NIGMS (5R35GM125034), and NIDDK (5U01DK112217)

Structural classification of peptide/HLA complexes enables accurate high-throughput modeling

Abstract The class I major histocompatibility complex (MHC-I) proteins display 8 to 15 residue peptides derived from endogenous proteins on the cell surface for immune surveillance. Accurate structural modeling of peptides bound to HLA (the human MHC) proteins has been mired by the inability to capture the central residues of the peptide. Here, an analysis of high-resolution X-ray crystal structures revealed that diverse peptide conformations arise as a result of local changes in the backbone configuration. As a result, we established an internal-coordinate based system to assess structural similarity between peptide backbones and found that peptide/HLA complexes, encompassing a wide range of allotypes and peptide sequences, present a discrete set of peptide backbone conformations. Leveraging these representative backbones and a regression model trained on a physically realistic scoring function, we developed RepPred, a structural modeling approach which predicts near native nonamer/HLA-A*02:01 complexes with high accuracy and throughput. Our method significantly outperformed five state-of-the-art peptide/HLA modeling approaches on the basis of structural accuracy. Thus, the structural information obtained from RepPred can be used to address long-standing immunology challenges such as predicting peptide immunogenicity. Supported by grants from NIH (R01AI143997, R35GM125034, U01DK112217).

Polarity induced vapochromism and vapoluminescence of polythiophene derivatives for volatile organic compounds classification

Polarity induced vapochromic and vapoluminescent properties of cationic poly-3-alkoxythiophene derivatives (PT) casted on polyvinylidene fluoride (PVDF) membranes are reported. PT with six different pendant groups are designed to differentially interact with volatile organic compounds (VOC) of varying polarities, thereby enabling their classification. PT exhibit a rapid vapochromic response with a concurrent modulation of vapoluminescence due to the non-covalent cation-π interactions between the pendant groups and the PT backbone. Adsorption of VOC on pendant groups alters the conformation of PT backbone, thus resulting in an increase in intensity and blue shifting of fluorescence emission within the visible spectrum. The vapoluminescent responses are found to be more sensitive with a limit of detection (LOD) of ∼7 ppm and a wider dynamic range as compared to the vapochromic responses with a LOD of ∼60 ppm for the detection of a model VOC: chloroform. Notably, all the PT illustrate an instantaneous recovery of colour and luminescence upon desorption of VOC. PT interaction with VOC of varying polarities was ascertained using density functional theory (DFT) and principal component analysis (PCA) methodologies. In summary, the polarity induced vapochromic and vapoluminescent properties of PT could yield a selective and sensitive vapochromic and fluorometric dual-mode VOC detection platform.

FLEXR: automated multi-conformer model building using electron-density map sampling.

Protein conformational dynamics that may inform biology often lie dormant in high-resolution electron-density maps. While an estimated ∼18% of side chains in high-resolution models contain alternative conformations, these are underrepresented in current PDB models due to difficulties in manually detecting, building and inspecting alternative conformers. To overcome this challenge, we developed an automated multi-conformer modeling program, FLEXR. Using Ringer-based electron-density sampling, FLEXR builds explicit multi-conformer models for refinement. Thereby, it bridges the gap of detecting hidden alternate states in electron-density maps and including them in structural models for refinement, inspection and deposition. Using a series of high-quality crystal structures (0.8-1.85 Å resolution), we show that the multi-conformer models produced by FLEXR uncover new insights that are missing in models built either manually or using current tools. Specifically, FLEXR models revealed hidden side chains and backbone conformations in ligand-binding sites that may redefine protein-ligand binding mechanisms. Ultimately, the tool facilitates crystallographers with opportunities to include explicit multi-conformer states in their high-resolution crystallographic models. One key advantage is that such models may better reflect interesting higher energy features in electron-density maps that are rarely consulted by the community at large, which can then be productively used for ligand discovery downstream. FLEXR is open source and publicly available on GitHub at https://github.com/TheFischerLab/FLEXR.

Connectivity between surface and interior in catalytic subunits of acetylcholinesterases inferred from their X-ray structures.

Catalytic activity and function of acetylcholinesterase (AChE; EC 3.1.1.7) have been recognized and studied for over a century and its quaternary and primary structures for about half a century, and its tertiary structure has been known for about 33 years. Clear understanding of relationships between the structure and the function is still pending for this enzyme. Hundreds of crystallographic, static snapshots of AChEs from different sources reveal largely one general backbone conformation with narrow entry into the active center gorge, tightly fit to accept one acetylcholine (ACh) molecule, in contrast to its high catalytic turnover. This short review of available X-ray structures of AChEs from electric ray Torpedo californica, mouse and human, finds some limited, yet consistent deviations in conformations of selected secondary structure elements of AChE relevant for its function. The observed conformational diversity of the acyl pocket loop of AChE, unlike the large Ω-loop, appears consistent with structurally dynamic INS data and solution-based SAXS experiments to explain its dominant role in controlling the size of the active center gorge opening, as well as connectivity between the immediate surroundings of the buried active Ser, and catalytically relevant sites on the AChE surface.