Well-defined Three-dimensional Structures Research Articles

Scalable Synthesis of All Stereoisomers of 2-Aminocyclopentanecarboxylic Acid─A Toolbox for Peptide Foldamer Chemistry.

Although the construction of peptides with well-defined three-dimensional structures and predictable functions, including biological activity, using conformationally constrained β-amino acids has been shown to be a very successful strategy, their broad application is limited by access to the appropriate building blocks. In particular, trans- and cis-stereoisomers of 2-aminocyclopentanecarboxylic acid (ACPC) are of high interest. The scalable synthesis of all four stereoisomers of Fmoc derivatives of ACPC is presented with NMR-based analysis methods for their enantiomeric purity.

Direct assembly of N-sulfamoyl lactam scaffolds bearing a zinc-binding group for inhibiting metalloenzymes based on desymmetrization of sulfamide and the Castagnoli-Cushman reaction

Sulfamide was desymmetrized by a reaction with aldehydes to give N-sulfamoylimines. The latter reagents were successfully introduced into diastereo- and chemoselective transformation with cyclic anhydride using the Castagnoli-Cushman reaction to give unprotected N-sulfamoyl tetrahydroisoquinolonic (THIQ) acids under simple metal-free protocol. Thereby, the first general approach to the direct assembly of six-membered N-sulfamoyl lactams was developed. The synthesized compounds belong to representative, drug-like chemotypes with their well-defined three-dimensional structures and tunable physicochemical properties. In an attempt to probe their pharmacological potential, the newly synthesized lactams were screened against therapeutically relevant human and bacterial carbonic anhydrases, with some derivatives showing low micromolar enzyme inhibitory profiles.

Spiroligomer-Based Macrocycles for Atomically Precise Membranes.

Here, we report a new class of peptidomimetic macrocycles with well-defined three-dimensional structures and low conformational flexibility. They are assembled from fused-ring spiro-ladder oligomers (spiroligomers) by modular solid-phase synthesis. Two-dimensional nuclear magnetic resonance confirms their shape persistency. Triangular macrocycles of tunable sizes assemble into membranes with atomically precise pores, which exhibit size and shape-dependent molecular sieving towards a series of structurally similar compounds. The exceptional structural diversity and stability of spiroligomer-based macrocycles will be explored for more applications.

CuO metallic aerogels with a tailored nodular morphology.

This work reports, for the first time, an efficient and fast microwave-based method for the preparation of CuO aerogels. For that, CuCl2, glyoxylic acid and sodium carbonate are employed as reagents. Different experimental conditions such as synthesis temperature, synthesis time and concentration of the precursor solution are investigated to design CuO aerogels with customizable nodular morphologies. The resulting aerogels exhibit well-defined three-dimensional structures and nodular sizes, and therefore, textural properties vary according to the experimental parameters applied in their synthesis.

Structural and Electronic Modulations of Imidazolium Covalent Organic Framework-Derived Electrocatalysts for Oxygen Redox Reactions in Rechargeable Zn-Air Batteries.

Covalent organic frameworks (COFs) are promising candidates for the controllable design of electrocatalysts. However, bifunctional electrocatalytic activities for the oxygen reduction reaction (ORR) and oxygen evolution reaction (OER) remain challenging in COFs. In this study, imidazolium-rich COFs (IMCOFs) with well-defined active sites and characteristic three-dimensional assembly structures were readily prepared, and their electronic structures were tuned by Co incorporation to elicit bifunctional electrocatalytic activities for the ORR and OER. The Co nanoparticle-incorporated spherical IMCOF-derived electrocatalyst (CoNP-s-IMCOF) exhibited lower overpotentials for the ORR and OER compared with the atomic Co-incorporated planar IMCOF-derived electrocatalyst (Co-p-IMCOF). Computational simulations revealed that the imidazole carbon sites of CoNP-s-IMCOF rather than the triazine carbons were the active sites for the ORR and OER, and its p-band center downshifted via charge transfer, facilitating the chemisorption of oxygen intermediates during the reactions. A Zn-air battery with CoNP-s-IMCOF exhibited a small voltage gap of 1.3 V with excellent durability for 935 cycles. This approach for control over the three-dimensional assembly and electronic structures of IMCOFs can be extended to the development of diverse catalytic nanomaterials for applications of interest.

Length-Dependent Collective Vibrational Dynamics in Alpha-Helices.

Functions of protein molecules in nature are closely associated with their well-defined three-dimensional structures and dynamics in body fluid. So far, many efforts have been made to reveal the relation of protein structure, dynamics, and function, but the structural origin of protein dynamics, especially for secondary structures as building blocks of conformation transition, is still ambiguous. Here we theoretically uncover the collective vibrations of elastic poly-alanine α-helices and find vibration patterns that are distinctively different over residue numbers ranging from 20 to 80. Contrary to the decreasing vibration magnitude from ends to the middle region for short helices, the vibration magnitude for long helices takes the minimum at approximately 1/5 of helix length from ends but reaches a peak at the center. Further analysis indicates that major vibrational modes of helical structures strongly depend on their lengths, where the twist mode dominates in the vibrations of short helices while the bend mode dominates the long ones analogous to an elastic Euler beam. The helix-coil transition pathway is also affected by the alternation of the first-order mode in helices with different lengths. The dynamic properties of the helical polypeptides are promising to be harnessed for de novo design of protein-based materials and artificial biomolecules in clinical treatments.

Chromophoric Dendrimer-Based Materials: An Overview of Holistic-Integrated Molecular Systems for Fluorescence Resonance Energy Transfer (FRET) Phenomenon.

Dendrimers (from the Greek dendros → tree; meros → part) are macromolecules with well-defined three-dimensional and tree-like structures. Remarkably, this hyperbranched architecture is one of the most ubiquitous, prolific, and recognizable natural patterns observed in nature. The rational design and the synthesis of highly functionalized architectures have been motivated by the need to mimic synthetic and natural-light-induced energy processes. Dendrimers offer an attractive material scaffold to generate innovative, technological, and functional materials because they provide a high amount of peripherally functional groups and void nanoreservoirs. Therefore, dendrimers emerge as excellent candidates since they can play a highly relevant role as unimolecular reactors at the nanoscale, acting as versatile and sophisticated entities. In particular, they can play a key role in the properties of light-energy harvesting and non-radiative energy transfer, allowing them to function as a whole unit. Remarkably, it is possible to promote the occurrence of the FRET phenomenon to concentrate the absorbed energy in photoactive centers. Finally, we think an in-depth understanding of this mechanism allows for diverse and prolific technological applications, such as imaging, biomedical therapy, and the conversion and storage of light energy, among others.

Nanoparticle conjugated with aptamer anti-MUC1/Y for inflammatory arthritis

Aptamers may form well-defined three-dimensional structures binding with high affinity and stability to a specific receptor. The aptamer anti-MUC1 isoform Y is one the most used due the affinity to MUC1, which is overexpressed in several types of cancer and inflammation process. In this study we have developed, characterized, in vitro as in vivo evaluated a nanoaptamer (anti-MUC1/Y) as a nanoagent for rheumatoid arthritis treatment. The results showed that a nanoaptamer with a size range of 241 nm was produced. The entrapment efficacy was 90% with a biodistribution showing a high hepatic uptake (>98%). The results in vivo showed a potent effect in arthritis experimental model, especially in low doses. The results corroborate the applicability of this nanosystem for RA treatment.

Fluorescence Depolarization Kinetics Captures Short-Range Backbone Dihedral Rotations and Long-Range Correlated Dynamics of an Intrinsically Disordered Protein.

Intrinsically disordered proteins (IDPs) do not autonomously fold into well-defined three-dimensional structures and are best described as a heterogeneous ensemble of rapidly interconverting conformers. It is challenging to elucidate their complex dynamic signatures using a single technique. In this study, we employed sensitive fluorescence depolarization kinetics by following picosecond time-resolved fluorescence anisotropy decays to directly capture the essential dynamical features of intrinsically disordered α-synuclein (α-syn) site-specifically labeled with thiol-active fluorophores. By utilizing a long-lifetime (≥10 ns) anisotropic label, we were able to discern three distinct rotational components of α-syn. The subnanosecond component represents the local wobbling-in-cone motion of the fluorophore, whereas the slower (∼1.4 ns) component corresponds to the short-range backbone dynamics governed by collective torsional fluctuations in the Ramachandran Φ-Ψ dihedral space. This backbone dihedral rotational time scale is sensitive to the local chain stiffness and slows down in the presence of an adjacent proline residue. We also observed a small-amplitude (≤10%) slower rotational correlation time (6-10 ns) that represents the long-range correlated dynamics involving a much longer segment of the polypeptide chain. These intrinsic dynamic signatures of IDPs will provide critical mechanistic underpinnings in a mosaic of biophysical phenomena involving internal friction, allosteric interactions, and phase separation.

Self-assembling protein nanocages for modular enzyme assembly by orthogonal bioconjugation.

The wide variety of enzymatic pathways that can benefit from enzyme scaffolding is astronomical. While enzyme co-localization based on protein, DNA, and RNA scaffolds has been reported, we still lack scaffolds that offer well-defined and uniform three-dimensional structures for enzyme organization. Here we reported a new approach for protein co-localization using naturally occurring protein nanocages as a scaffold. Two different nanocages, the 25 nm E2 and the 34 nm heptatitis B virus, were used to demonstrate the successfully co-localization of the endoglucanase CelA and cellulose binding domain using the robust SpyTag/SpyCatcher bioconjugation chemistry. Because of the simplicity of the assembly, this strategy is useful not only for in vivo enzyme cascading but also the potential for in vivo applications as well.

Determination of minimal sequence for zearalenone aptamer by computational docking and application on an indirect competitive electrochemical aptasensor.

Aptamers are short single-stranded oligonucleotides (either DNA or RNA) that can fold into well-defined three-dimensional (3D) spatial structures which enable them to capture their specific target by complementary shape interactions. Aptamers are selected from large random libraries through the SELEX process and only a small fraction of the sequence is involved in direct docking with the target. In this paper, we describe the possible truncation variants of zearalenone (ZEA) aptamer which might be an effective binding region for the target. The originally selected zearalenone (ZEA) aptamer was 80-mer in length and shown to bind the target with a high affinity (Kd = 41 ± 5nM). Herein, computational docking simulation was performed with 15 truncated variants to determine the predicted binding energy and responsible binding site of the aptamer-analyte complex. The results revealed that 5 truncated variants had binding energy lower than - 7.0kcal/mol. Circular dichroism analysis was performed on the shortlisted aptamer and the conformational change of aptamers was observed with the presence of an analyte. Aptamer Z3IN (29-mer) was chosen as the most enhanced affinity for its target with a dissociation constant of 11.77 ± 1.44nM. The aptamer was further applied in the electrochemical aptasensor of ZEA based on an indirect competitive format. The results demonstrated that the truncated aptamer leads to an enhancement of the sensitivity of the biosensor.

Exploring the conformational dynamics and flexibility of intrinsically disordered HIV-1 Nef protein using molecular dynamic network approaches.

The online version contains supplementary material available at 10.1007/s13205-021-02698-8.

Green Synthesis of Free Standing Cellulose/Graphene Oxide/Polyaniline Aerogel Electrode for High-Performance Flexible All-Solid-State Supercapacitors.

The cellulose/graphene oxide (GO) networks as the scaffold of free-standing aerogel electrodes are developed by using lithium bromide aqueous solution, as the solvent, to ensure the complete dissolution of cotton linter pulp and well dispersion/reduction of GO nanosheets. Polyaniline (PANI) nanoclusters are then coated onto cellulose/GO networks via in-situ polymerization of aniline monomers. By optimized weight ratio of GO and PANI, the ternary cellulose/GO3.5/PANI aerogel film exhibits well-defined three-dimensional porous structures and high conductivity of 1.15 S/cm, which contributes to its high areal specific capacitance of 1218 mF/cm2 at the current density of 1.0 mA/cm2. Utilizing this cellulose/GO3.5/PANI aerogel film as electrodes in a symmetric configuration supercapacitor can result in an outstanding energy density as high as 258.2 µWh/cm2 at a power density of 1201.4 µW/cm2. Moreover, the device can maintain nearly constant capacitance under different bending deformations, suggesting its promising applications in flexible electronics.

Secondary Structure Prediction for Intrinsically Disordered Proteins

Intrinsically disordered proteins (IDPs) or intrinsically disordered regions (IDRs), without well-defined three-dimensional structures, play vital roles in various biological processes, e.g., protein-/nucleic acid-binding, cellular signaling, and protein regulation. One such role of IDPs/IDRs is the intracellular compartmentalization by forming membraneless organelles via liquid-liquid phase separation (LLPS). These phase-separated protein assemblies are also vital in the design of biomimetic protein-based materials. We recently developed a residue-level coarse-grained (CG) model that has yielded valuable information regarding LLPS of various IDPs [1-2]. However, the current model treats protein chains as flexible molecules in which harmonic bonds link amino acid interaction sites. The model lacks chain rigidity and the ability to populate common secondary structure elements due to the lack of angle and dihedral backbone potentials. Here, we present a new CG model which includes these additional backbone potentials to help capture the transient secondary structures of IDPs. The parameters for the pseudo backbone dihedral potential are transferable and obtained from the context-independent helical propensities of twenty naturally occurring amino acids using NMR [3]. We show that the new model can accurately predict the regions of high helical propensities of IDPs such as TDP-43 as well as PPII propensities of proline-rich regions when compared to NMR and all-atom simulation data. Looking forward, this model can be an essential tool in understanding the sequence-specific LLPS of IDPs and high-throughput sequence protein design. 1. G. L. Dignon, W. Zheng, R. B. Best, Y. C. Kim, and J. Mittal, Proc. Nat. Acad. Sci. 115, 9929-9934, 2018. 2. G. L. Dignon, W. Zheng, Y. C. Kim, and J. Mittal, ACS Cent. Sci. 5, 821-830, 2019. 3. R. J. Moreau, et al., J. Am. Chem. Soc. 131, 13107-13116, 2009.

Rational De Novo Design of a Cu Metalloenzyme for Superoxide Dismutation.

Superoxide dismutases (SODs) are highly efficient enzymes for superoxide dismutation and the first line of defense against oxidative stress. These metalloproteins contain a redox-active metal ion in their active site (Mn, Cu, Fe, Ni) with a tightly controlled reduction potential found in a close range around the optimal value of 0.36 V versus the normal hydrogen electrode (NHE). Rationally designed proteins with well-defined three-dimensional structures offer new opportunities for obtaining functional SOD mimics. Here, we explore four different copper-binding scaffolds: H3 (His3 ), H4 (His4 ), H2 DH (His3 Asp with two His and one Asp in the same plane) and H3 D (His3 Asp with three His in the same plane) by using the scaffold of the de novo protein GRα3 D. EPR and XAS analysis of the resulting copper complexes demonstrates that they are good CuII -bound structural mimics of Cu-only SODs. Furthermore, all the complexes exhibit SOD activity, though three orders of magnitude slower than the native enzyme, making them the first de novo copper SOD mimics.

Features of molecular recognition of intrinsically disordered proteins via coupled folding and binding.

The sequence-structure-function paradigm of proteins has been revolutionized by the discovery of intrinsically disordered proteins (IDPs) or intrinsically disordered regions (IDRs). In contrast to traditional ordered proteins, IDPs/IDRs are unstructured under physiological conditions. The absence of well-defined three-dimensional structures in the free state of IDPs/IDRs is fundamental to their function. Folding upon binding is an important mode of molecular recognition for IDPs/IDRs. While great efforts have been devoted to investigating the complex structures and binding kinetics and affinities, our knowledge on the binding mechanisms of IDPs/IDRs remains very limited. Here, we review recent advances on the binding mechanisms of IDPs/IDRs. The structures and kinetic parameters of IDPs/IDRs can vary greatly, and the binding mechanisms can be highly dependent on the structural properties of IDPs/IDRs. IDPs/IDRs can employ various combinations of conformational selection and induced fit in a binding process, which can be templated by the target and/or encoded by the IDP/IDR. Further studies should provide deeper insights into the molecular recognition of IDPs/IDRs and enable the rational design of IDP/IDR binding mechanisms in the future.

Ubiquitin Designer Proteins as a New Additive Generation toward Controlling Crystallization.

Proteins controlling mineralization in vivo are diverse, suggesting that there are various ways by which mineralization can be directed in bioinspired approaches. While well-defined three-dimensional (3D) structures occur in biomineralization proteins, the design of synthetic, soluble, bioinspired macromolecules with specific, reproducible, and predictable 3D arrangements of mineral-interacting functions poses an ultimate challenge. Thus, the question of how certain arrangements of such functions on protein surfaces influence mineralization and in what ways specific alterations subsequently affect this process remains elusive. Here we used genetically engineered ubiquitin (Ub) proteins in order to overcome the limitations of generic bioinspired additive systems. By advancing existing protocols, we introduced an unnatural amino acid and subsequently mineral-interacting functions via selective-pressure incorporation and click chemistry, respectively, without affecting the Ub secondary structure. Indeed, as-obtained Ub with three phosphate functions at defined positions shows unique effects based on a yet-unmatched capability toward the stabilization of a film of a dense liquid mineral phase visible even with the naked eye and its transformation into amorphous nanoparticles and afterward crystals with complex shapes. We thereby demonstrate that Ub designer proteins pose a unique new generation of crystallization additives where the 3D arrangement of mineral-interacting functions can be designed at will, promising their future use for programmable, target-oriented mineralization control.

Sequence Effects on Size, Shape, and Structural Heterogeneity in Intrinsically Disordered Proteins.

Intrinsically disordered proteins (IDPs) lack well-defined three-dimensional structures, thus challenging the archetypal notion of structure-function relationships. Determining the ensemble of conformations that IDPs explore under physiological conditions is the first step toward understanding their diverse cellular functions. Here, we quantitatively characterize the structural features of IDPs as a function of sequence and length using coarse-grained simulations. For diverse IDP sequences, with the number of residues ( NT) ranging from 20 to 441, our simulations not only reproduce the radii of gyration ( Rg) obtained from experiments, but also predict the full scattering intensity profiles in excellent agreement with small-angle X-ray scattering experiments. The Rg values are well-described by the standard Flory scaling law, Rg = Rg0 NTν, with ν ≈ 0.588, making it tempting to assert that IDPs behave as polymers in a good solvent. However, clustering analysis reveals that the menagerie of structures explored by IDPs is diverse, with the extent of heterogeneity being highly sequence-dependent, even though ensemble-averaged properties, such as the dependence of Rg on chain length, may suggest synthetic polymer-like behavior in a good solvent. For example, we show that for the highly charged Prothymosin-α, a substantial fraction of conformations is highly compact. Even if the sequence compositions are similar, as is the case for α-Synuclein and a truncated construct from the Tau protein, there are substantial differences in the conformational heterogeneity. Taken together, these observations imply that metrics based on net charge or related quantities alone cannot be used to anticipate the phases of IDPs, either in isolation or in complex with partner IDPs or RNA. Our work sets the stage for probing the interactions of IDPs with each other, with folded protein domains, or with partner RNAs, which are critical for describing the structures of stress granules and biomolecular condensates with important cellular functions.

Functionalized Helical β-Peptoids.

Peptidomimetic foldamers adopting well-defined three-dimensional structures while being stable toward proteolysis are of interest in biomedical research, chemical biology, and biomimetic materials science. Despite their backbone flexibility, β-peptoids containing N-( S)-1-(1-naphthyl)ethyl ( Ns1npe) side chains can fold into unique triangular prism-shaped helices. We report herein the successful introduction of amino groups onto robustly folded β-peptoid helices by construction and incorporation of novel chiral building blocks. This is the first example of an X-ray crystal structure of a linear β-peptoid containing more than one type of side chain. We thus present a unique foldamer design comprising a robustly folded core with functionalized side chains protruding perpendicular to the helical axis to provide a highly predictable display of functional groups. This work paves the way for development of β-peptoid foldamers with a desired function, such as catalytic properties or as scaffolds enabling polyvalent display.

Calcareous Foraminiferal Shells as a Template for the Formation of Hierarchal Structures of Inorganic Nanomaterials

A microorganism template approach has been explored for the fabrication of various well-defined three-dimensional (3D) structures. However, most of these templates suffer from small size (few μm), difficulty to remove the template, or low surface area, which affect their potential use in different applications or makes industrial scale-up difficult. Conversely, foraminifer's microorganisms are large (up to 200 mm), consist of CaCO3 (easy to dissolve in mild acid), and have a relatively high surface area (≈5 m2 g-1). Herein, we demonstrate the formation of hierarchical structures of inorganic materials using calcareous foraminiferal shells such as Sorites, Globigerinella siphonifera, Lox-ostomina amygdaleformis, Calcarina baculatus or hispida, and Peneroplis planatus. Several techniques, such as thermal decomposition of single-source precursors of metal oxides or sulfides, reduction of metal salts directly on the surfaces, and redox reactions, were used for coating of different shell materials and several hybrid compositions, which possess nanofeatures. Finally, we examined the role of the prepared 3D structures on the reduction of 4-nitrophenol (4-NP), ethanol electrooxidation, and water purification. A remarkable performance was achieved in each application. The hierarchical structure leads to the reduction of 4-NP within several minutes, a 27 mA cm-2 current density peak was obtained for ethanol electrooxidation, and more than 95% of the organic dye contaminants were successfully removed. These results show that using foraminiferal shells offers a new way for designing complex hierarchical structures with unique properties.