Peptides are naturally potent and selective therapeutics with massive potential; however, low cell membrane permeability limits their clinical implementation, particularly for hydrophilic, anionic peptides with intracellular targets. To overcome this limitation, esterification of anionic carboxylic acids on therapeutic peptides can simultaneously increase hydrophobicity and net charge to facilitate cell internalization, whereafter installed esters can be cleaved hydrolytically to restore activity. To date, however, most esterified therapeutics contain either a single esterification site or multiple esters randomly incorporated on multiple sites. This investigation provides molecular engineering insight into how the number and position of esters installed onto the therapeutic peptide α carboxyl terminus 11 (αCT11, RPRPDDLEI) with 4 esterification sites affect hydrophobicity and the hydrolysis process that reverts the peptide to its original form. After installing methyl esters onto αCT11 using Fischer esterification, we isolated 5 distinct products and used 2D nuclear magnetic resonance spectroscopy, reverse-phase high performance liquid chromatography, and mass spectrometry to determine which residues were esterified in each and the resulting increase in hydrophobicity. We found esterifying the C-terminal isoleucine to impart the largest increase in hydrophobicity. Monitoring ester hydrolysis showed the C-terminal isoleucine ester to be the most hydrolytically stable, followed by the glutamic acid, whereas esters on aspartic acids hydrolyze rapidly. LC-MS revealed the formation of transient intramolecular aspartimides prior to hydrolysis to carboxylic acids. proof-of-concept experiments showed esterifying αCT11 to increase cell migration into a scratch, highlighting the potential of multi-site esterification as a tunable, reversible strategy to enable the delivery of therapeutic peptides.

3D Voronoi scaffolds are widely applied in the field of additive manufacturing as they are known for their light weight structural resilience and share many topological similarities to various natural (bone, tumours, lymph node) and synthetic environments (foam, functionally gradient porous materials). Unfortunately, the structural design features that promote these topological similarities (such as the number of vertices) are often unpredictable and require the trial and error of varying design features to achieve the desired 3D Voronoi structure. This article provides a toolkit, consisting of equations, based on over 12 000 3D Voronoi structures. These equations allow design features, such as the number of generating points (), to be efficiently and accurately predicted based on the desired structural parameters (within ±3). Based on these equations we are proposing, to the best of our knowledge, two new mathematical conjectures that relate the number of vertices or edges, and the average edge length to in Voronoi structures. These equations have been validated for a wide range of parameter values and Voronoi network sizes. A design code is provided allowing any of over 12 000 structures to be selected, easily adjusted based on user requirements, and 3D printed. Biomedical case studies relevant to T-cell culturing, bone scaffolds and kidney tumours are presented to illustrate the design code.

The molecular-level understanding of intrinsically disordered proteins is challenging due to experimental characterization difficulties. Computational understanding of IDPs also requires fundamental advances, as the leading tools for predicting protein folding (e.g., AlphaFold), typically fail to describe the structural ensembles of IDPs. The focus of this paper is to 1) develop new representations for intrinsically disordered proteins and 2) pair these representations with classical machine learning and deep learning models to predict the radius of gyration and derived scaling exponent of IDPs. Here, we build a new physically-motivated feature called the bag of amino acid interactions representation, which encodes pairwise interactions explicitly into the representation. This feature essentially counts and weights all possible non-bonded interactions in a sequence and thus is, in principle, compatible with arbitrary sequence lengths. To see how well this new feature performs, both categorical and physically-motivated featurization techniques are tested on a computational dataset containing 10,000 sequences simulated at the coarse-grained level. The results indicate that this new feature outperforms the other purely categorical and physically-motivated features and possesses solid extrapolation capabilities. For future use, this feature can potentially provide physical insights into amino acid interactions, including their temperature dependence, and be applied to other protein spaces.

We present a method of enabling photochemical reactions in water by using biomimetic, water-soluble liposomes and a specifically functionalized perylene diimide chromophore. Linking two flexible saturated C4-alkyl chains with terminal positively charged trimethylammonium groups to the rigid perylene diimide core yielded 1 allowing for its co-assembly at the lipid bilayer interface of DOPG liposomes (DOPG = 1,2-dioleoyl--3-phospho-(1'--glycerol)) with a preferred orientation and in close proximity to the water interface. According to molecular dynamics simulations the chromophore aligns preferably parallel to the membrane surface which is supported by confocal microscopy. Irradiation experiments with visible light and in the presence of a negatively charged, water-soluble oxidant were slower in the DOPG-membrane than under acetonitrile-water reaction conditions. The generated radical species was characterized by EPR spectroscopy in an acetonitrile-water mixture and associated to the DOPG-membrane. Time-resolved emission studies revealed a static quenching process for the initial electron transfer from photoexcited 1 to the water soluble oxidant. The findings presented in this study yield design principles for the functionalization of lipid bilayer membranes which will be relevant for the molecular engineering of artificial cellular organelles and nano-reactors based on biomimetic vesicles and membranes.

Conjugated macrocycles can exhibit concealed antiaromaticity; that is, despite not being antiaromatic, under specific circumstances, they can display properties typically observed in antiaromatic molecules due to their formal macrocyclic 4 π-electron system. Paracyclophanetetraene (PCT) and its derivatives are prime examples of macrocycles exhibiting this behaviour. In redox reactions and upon photoexcitation, they have been shown to behave like antiaromatic molecules (requiring type I and II concealed antiaromaticity, respectively), with such phenomena showing potential for use in battery electrode materials and other electronic applications. However, further exploration of PCTs has been hindered by the lack of halogenated molecular building blocks that would permit their integration into larger conjugated molecules by cross-coupling reactions. Here, we present two dibrominated PCTs, obtained as a mixture of regioisomers from a three-step synthesis, and demonstrate their functionalisation Suzuki cross-coupling reactions. Optical, electrochemical, and theoretical studies reveal that aryl substituents can subtly tune the properties and behaviour of PCT, showing that this is a viable strategy in further exploring this promising class of materials.

Due to their N-substitution, peptoids are generally regarded as resistant to biological degradation, such as enzymatic and hydrolytic mechanisms. This stability is an especially attractive feature for therapeutic development and is a selling point of many previous biological studies. However, another key mode of degradation remains to be fully explored, namely oxidative degradation mediated by reactive oxygen and nitrogen species (ROS/RNS). ROS and RNS are biologically relevant in numerous contexts where biomaterials may be present, thus, improving understanding of peptoid oxidative susceptibility is crucial to exploit their full potential in the biomaterials field, where an oxidatively-labile but enzymatically stable molecule can offer attractive properties. Toward this end, we demonstrate a fundamental characterization of sequence-defined peptoid chains in the presence of chemically generated ROS, as compared to ROS-susceptible peptides such as proline and lysine oligomers. Lysine oligomers showed the fastest degradation rates to ROS and the enzyme trypsin. Peptoids degraded in metal catalyzed oxidation conditions at rates on par with poly(prolines), while maintaining resistance to enzymatic degradation. Furthermore, lysine-containing peptide-peptoid hybrid molecules showed tunability in both ROS-mediated and enzyme-mediated degradation, with rates intermediate to lysine and peptoid oligomers. When lysine-mimetic side-chains were incorporated into a peptoid backbone, the rate of degradation matched that of the lysine peptide oligomers, but remained resistant to enzymatic degradation. These results expand understanding of peptoid degradation to oxidative and enzymatic mechanisms, and demonstrate the potential for peptoid incorporation into materials where selectivity towards oxidative degradation is necessary, or directed enzymatic susceptibility is desired.

Herein, we report the crystal structure of 2,7-dichlorofluorescein methyl ester (DCF-ME) and its fluorescence response to hydroxyapatite binding. The reported fluorophore is very selective for staining the bone matrix and provides turn-on fluorescence upon hydroxyapatite binding. The reported fluorophore can readily pass the cell membrane of the C2C12 cell line, and it is non-toxic for the cell line. The reported fluorophore DCF-ME may find applications in monitoring bone remodeling and microcalcification as an early diagnosis tool for breast cancer and age-related macular degeneration.

The ease of functionalization of metal-organic frameworks (MOFs) can unlock unprecedented opportunities for gas adsorption and separation applications as the functional groups can impart favorable/unfavorable regions/interactions for the desired/undesired adsorbates. In this study, the effects of the presence of multiple functional groups in MOFs on their CF/CH, CH/H, CH/N, and N/H separation performances were computationally investigated combining grand canonical Monte Carlo (GCMC) and molecular dynamics (MD) simulations. The most promising adsorbents showing the best combinations of selectivity, working capacity, and regenerability were identified for each gas separation. 15, 13, and 16 out of the top 20 MOFs identified for the CH/H, CH/N, and N/H adsorption-based separation, respectively, were found to have -OCH groups as one of the functional groups. The biggest improvements in CF/CH, CH/H, CH/N, and N/H selectivities were found to be induced by the presence of -OCH-OCH groups in MOFs. For CH/H separation, MOFs with two and three functionalized linkers were the best adsorbent candidates while for N/H separation, all the top 20 materials involve two functional groups. Membrane performances of the MOFs were also studied for CH/H and CH/N separation and the results showed that MOFs having -F-NH and -F-OCH functional groups present the highest separation performances considering both the membrane selectivity and permeability.

Labeled protein-based biomaterials have become a popular for various biomedical applications such as tissue-engineered, therapeutic, or diagnostic scaffolds. Labeling of protein biomaterials, including with ultrasmall super-paramagnetic iron oxide (USPIO) nanoparticles, has enabled a wide variety of imaging techniques. These USPIO-based biomaterials are widely studied in magnetic resonance imaging (MRI), thermotherapy, and magnetically-driven drug delivery which provide a method for direct and non-invasive monitoring of implants or drug delivery agents. Where most developments have been made using polymers or collagen hydrogels, shown here is the use of a rationally designed protein as the building block for a meso-scale fiber. While USPIOs have been chemically conjugated to antibodies, glycoproteins, and tissue-engineered scaffolds for targeting or improved biocompatibility and stability, these constructs have predominantly served as diagnostic agents and often involve harsh conditions for USPIO synthesis. Here, we present an engineered protein-iron oxide hybrid material comprised of an azide-functionalized coiled-coil protein with small molecule binding capacity conjugated via bioorthogonal azide-alkyne cycloaddition to an alkyne-bearing iron oxide templating peptide, CMms6, for USPIO biomineralization under mild conditions. The coiled-coil protein, dubbed Q, has been previously shown to form nanofibers and, upon small molecule binding, further assembles into mesofibers via encapsulation and aggregation. The resulting hybrid material is capable of doxorubicin encapsulation as well as sensitive *-weighted MRI darkening for strong imaging capability that is uniquely derived from a coiled-coil protein.

Tuberculosis (TB) is an air-borne infectious disease and is the leading cause of death among all infectious diseases globally. The current treatment regimen for TB is overtly long and patient non-compliance often leads to drug resistant TB resulting in a need to develop new drugs that will act via novel mechanisms. In this research work, we selected membrane protein large (MmpL3) as the drug target and indole-2-carboximide as our molecule of interest for further designing new molecules. A homology model was prepared for the MmpL3 from the crystal structure of MmpL3. A series of indoles which are known to be MmpL3 inhibitors were docked in the prepared protein and the binding site properties were identified. Based on that, 10 molecules were designed and synthesized and their antitubercular activities evaluated. We identified four hits among which the highest potency candidate possessed a minimum inhibitory concentration (MIC) of 1.56 μM at 2-weeks. Finally, molecular dynamics simulation studies were done with 3b and a previously reported MmpL3 inhibitor to understand the intricacies of their binding in real time and to correlate the experimental findings with the simulation data.

For a series of phospholipid coated calamitic nematic liquid crystal droplets (5CB, 6CB, 7CB, E7 and MLC7023) of diameter ∼18 μm, the addition of chiral dopant leaves the sign of surface anchoring unchanged. Herein we report that for these chiral nematic droplets an analyte induced transition from a Frank-Pryce structure (with planar anchoring) to a nested-cup structure (with perpendicular anchoring) is accompanied by changes in the intensity of reflected light. We propose this system as both a general scheme for understanding director fields in chiral nematic liquid crystal droplets with perpendicular anchoring and as an ideal candidate to be utilised as the basis for developing cheap, single use LC-based sensor devices.

Decades of work in the field of computational study of semiconducting polymers using atomistic models illustrate the challenges of generating equilibrated models for this class of materials. While adopting a coarse-grained model can be helpful, the process of developing a suitable model is particularly non-trivial and time-consuming for semiconducting polymers due to a large number of different interactions with some having an anisotropic nature. This work introduces a procedure for the rapid generation of a hybrid model for semiconducting polymers where atoms of secondary importance (those in the alkyl side chains) are transformed into coarse-grained beads to reduce the computational cost of generating an equilibrated structure. The parameters are determined from easy-to-equilibrate simulations of very short oligomers and the model is constructed to enable a very simple back-mapping procedure to reconstruct geometries with atomistic resolution. The model is illustrated for three related polymers containing DPP (diketopyrrolopyrrole) to evaluate the transferability of the potential across different families of polymers. The accuracy of the model, determined by comparison with the results of fully equilibrated simulations of the same material before and after back-mapping, is fully satisfactory for two out of the three cases considered. We noticed that accuracy can be determined very early in the workflow so that it is easy to assess when the deployment of this method is advantageous. The hybrid representation can be used to evaluate directly the electronic properties of structures sampled by the simulations.

Although researchers have been working tirelessly since the COVID-19 outbreak, so far only three drugs - remdesivir, ronapreve and molnupiravir - have been approved for use in some countries which directly target the SARS-CoV-2 virus. Given the slow pace and substantial costs of new drug discovery and development, together with the urgency of the matter, repurposing of existing drugs for the ongoing disease is an attractive proposition. In a recent study, a high-throughput X-ray crystallographic screen was performed for a selection of drugs which have been approved or are in clinical trials. Thirty-seven compounds have been identified from drug libraries all of which bind to the SARS-CoV-2 main protease (3CL). In the current study, we use molecular dynamics simulation and an ensemble-based free energy approach, namely, enhanced sampling of molecular dynamics with approximation of continuum solvent (ESMACS), to investigate a subset of the aforementioned compounds. The drugs studied here are highly diverse, interacting with different binding sites and/or subsites of 3CL. The predicted free energies are compared with experimental results wherever they are available and they are found to be in excellent agreement. Our study also provides detailed energetic insights into the nature of the associated drug-protein binding, in turn shedding light on the design and discovery of potential drugs.

The negative membrane potential of bacterial cells influences crucial cellular processes. Inspired by the molecular scaffold of the antimicrobial peptide PGLa, we have developed antimicrobial foldamers with a computer-guided design strategy. The novel PGLa analogues induce sustained membrane hyperpolarization. When co-administered as an adjuvant, the resulting compounds - PGLb1 and PGLb2 - have substantially reduced the level of antibiotic resistance of multi-drug resistant , and clinical isolates. The observed antibiotic potentiation was mediated by hyperpolarization of the bacterial membrane caused by the alteration of cellular ion transport. Specifically, PGLb1 and PGLb2 are selective ionophores that enhance the Goldman-Hodgkin-Katz potential across the bacterial membrane. These findings indicate that manipulating bacterial membrane electrophysiology could be a valuable tool to overcome antimicrobial resistance.

In nature, the precise heterogeneous co-assembly of different protein domains gives rise to supramolecular machines that perform complex functions through the co-integrated activity of the individual protein subunits. A synthetic approach capable of mimicking this process would afford access to supramolecular machines with new or improved functional capabilities. Here we show that the distinct peptide strands of a heterotrimeric α-helical coiled-coil (i.e., peptides "A", "B", and "C") can be used as fusion tags for heterogeneous co-assembly of proteins into supramolecular structures with tunable subunit stoichiometry. In particular, we demonstrate that recombinant fusion of A with NanoLuc luciferase (NL-A), B with superfolder green fluorescent protein (sfGFP-B), and C with mRuby (mRuby-C) enables formation of ternary complexes capable of simultaneously emitting blue, green, and red light via sequential bioluminescence and fluorescence resonance energy transfer (BRET/FRET). Fusion of galectin-3 onto the C-terminus of NL-A, sfGFP-B, and mRuby-C endows the ternary complexes with lactose-binding affinity that can be tuned by varying the number of galectin-3 domains integrated into the complex from one to three, while maintaining BRET/FRET function. The modular nature of the fusion protein design, the precise control of domain stoichiometry, and the multiplicity afforded by the three-stranded coiled-coil scaffold provides access to a greater range of subunit combinations than what is possible with heterodimeric coiled-coils used previously. We envision that access to this expanded range of co-integrated protein domain diversity will be advantageous for future development of designer supramolecular machines for therapeutic, diagnostic, and biotechnology applications.

Inspired by Nature's tunability driven by the modulation of structural organization, we utilize peptide motifs as an approach to tailor not only hierarchical structure, but also thermo-responsive shape memory properties of conventional polymeric materials. Specifically, poly(β-benzyl-L-aspartate)--poly(dimethylsiloxane)--poly(β-benzyl-L-aspartate) was incorporated as the soft segment in peptide-polyurea hybrids to manipulate hierarchical ordering through peptide secondary structure and a balance of inter- and intra-molecular hydrogen bonding. Employing these bioinspired peptidic polyureas, we investigated the influence of secondary structure on microphase-separated morphology, and shape fixity and recovery attenuated total reflectance-Fourier transform infrared spectroscopy (ATR-FTIR), small-angle X-ray scattering (SAXS) and dynamic mechanical analysis (DMA). The β-sheet motifs promoted phase mixing through extensive inter-molecular hydrogen bonding between the hard block and peptide segments and provided an increased chain elasticity, resulting in decreased shape fixity compared to a non-peptidic control. In contrast, intra-molecular hydrogen bonding driven by the α-helical arrangements yielded a microphase-separated and hierarchically ordered morphology, leading to an increase in the shape fixing ratio. These results indicate that peptide secondary structure provides a convenient handle for tuning shape memory properties by regulating hydrogen bonding with the surrounding polyurea hard segment, wherein extent of hydrogen bonding and phase mixing between the peptidic block and hard segment dictate the resulting shape memory behaviour. Furthermore, the ability to shift secondary structure as a function of temperature was also demonstrated as a pathway to influence shape memory response. This research highlights that peptide secondary conformation influences the hierarchical ordering and modulates the shape memory response of peptide-polymer hybrids. We anticipate that these findings will enable the design of smart bio-inspired materials with responsive and tailored function a balance of hydrogen bonding character, structural organization, and mechanics.

The advancement of click-functionalized hydrogels in recent years has coincided with rapid growth in the fields of mechanobiology, tissue engineering, and regenerative medicine. Click chemistries represent a group of reactions that possess high reactivity and specificity, are cytocompatible, and generally proceed under physiologic conditions. Most notably, the high level of tunability afforded by these reactions enables the design of user-controlled and tissue-mimicking hydrogels in which the influence of important physical and biochemical cues on normal and aberrant cellular behaviors can be independently assessed. Several critical tissue properties, including stiffness, viscoelasticity, and biomolecule presentation, are known to regulate cell mechanobiology in the context of development, wound repair, and disease. However, many questions still remain about how the individual and combined effects of these instructive properties regulate the cellular and molecular mechanisms governing physiologic and pathologic processes. In this review, we discuss several click chemistries that have been adopted to design dynamic and instructive hydrogels for mechanobiology investigations. We also chart a path forward for how click hydrogels can help reveal important insights about complex tissue microenvironments.

Zirconium metal-organic frameworks (MOFs) can be promising adsorbents for various applications as they are highly stable in different chemical environments. In this work, a collection of Zr-MOFs comprised of more than 100 materials is screened for CF/CH, CH/H, and CH/N separations using atomistic-level simulations. The top three MOFs for the CF/CH separation are identified as PCN-700-BPDC-TPDC, LIFM-90, and BUT-67 exhibiting CF/CH adsorption selectivities of 4.8, 4.6, and 4.7, CF working capacities of 2.0, 2.0, and 2.1 mol kg, and regenerabilities of 85.1, 84.2, and 75.7%, respectively. For the CH/H separation, MOF-812, BUT-67, and BUT-66 are determined to be the top performing MOFs demonstrating CH/H selectivities of 61.6, 36.7, and 46.2, CH working capacities of 3.0, 4.1, and 3.4 mol kg, and CH regenerabilities of 70.7, 82.7, and 74.7%, respectively. Regarding the CH/N separation, BUT-67, Zr-AbBA, and PCN-702 achieving CH/N selectivities of 4.5, 3.4, and 3.8, CH working capacities of 3.6, 3.9, and 3.5 mol kg, and CH regenerabilities of 81.1, 84.0, and 84.5%, in successive order, show the best overall separation performances. To further elucidate the adsorption in top performing adsorbents, the adsorption sites in these materials are analyzed using radial distribution functions and adsorbate density profiles. Finally, the water affinities of Zr-MOFs are explored to comment on their practical use in real gas separation applications. Our findings may inspire future studies probing the adsorption/separation mechanisms and performances of Zr-MOFs for different gases.

Elastin-like polypeptides (ELP) have been widely used in the biomaterials community due to their controllable, thermoresponsive properties and biocompatibility. Motivated by our previous work on the effect of tryptophan (W) substitutions on the LCST-like transitions of short ELPs, we studied a series of short ELPs containing tyrosine (Y) and/or phenylalanine (F) guest residues with only 5 or 6 pentapeptide repeat units. A combination of experiments and molecular dynamics (MD) simulations illustrated that the substitution of F with Y guest residues impacted the transition temperature (T) of short ELPs when conjugated to collagen-like-peptides (CLP), with a reduction in the transition temperature observed only after substitution of at least two residues. Placement of the Y residues near the N-terminal end of the ELP, away from the tethering point to the CLP, resulted in a lower T than that observed for peptides with the Y residues near the tethering point. Atomistic and coarse-grained MD simulations indicated an increase in intra- and inter- peptide hydrogen bonds in systems containing Y guest residues that are suggested to enhance the ability of the peptides to coacervate, with a concomitantly lower T. Simulations also revealed that the placement of Y-containing pentads near the N-terminus (i.e., away from CLP tethering point) versus C-terminus of the ELP led to more π-π stacking interactions at low temperatures, in agreement with our experimental observations of a lower T. Overall, this study provides mechanistic insights into the driving forces for the LCST-like transitions of ELPs and offers additional means for tuning the T of short ELPs for biomedical applications such as on-demand drug delivery and tissue engineering.

One proposed mechanism of implant fouling is attributed to the nonspecific adsorption of non-collagenous bone matrix proteins (NCPs) onto a newly implanted interface. With the goal of capturing the fundamental mechanistic and thermodynamic forces that govern changes in these NCP recognition domains as a function of γ-carboxyglutamic acid (Gla) post-translational modification and surface chemistry, we probe the adsorption process of the most commonly occurring NCP, osteocalcin, onto a mineral and metal oxide surface. Here, we apply two enhanced sampling methods to independently probe the effects of post-translational modification and peptide structure on adsorption. First, well-tempered metadynamics was used to capture the binding of acetyl and N-methylamide capped glutamic acid and Gla single amino acids onto crystalline hydroxyapatite and titania model surfaces at physiological pH. Following this, parallel tempering metadynamics in the well-tempered ensemble (PTMetaD-WTE) was used to study adsorption of the α-1 domain of osteocalcin onto hydroxyapatite and titania. Simulations were performed for the α-1 domain of osteocalcin in both its fully decarboxylated (dOC) and fully carboxylated (OC) form. Our simulations find that increased charge density due to carboxylation results in increased interactions at the interface, and stronger adsorption of the single amino acids to both surfaces. Interestingly, the role of Gla in promoting compact and helical structure in the α-1 domain resulted in disparate binding modes at the two surfaces, which is attributed to differences in interfacial water behavior. Overall, this work provides a benchmark for understanding the mechanisms that drive adsorption of Gla-containing mineralizing proteins onto different surface chemistries.

A mixed-ligand metal-organic framework (MOF) material composed of both amine- and hydroxyl-bearing linkers, KSU-1, was reacted with a variety of isocyanates. The hydroxyl groups reacted to a greater extent than the amines, in conflict with the previously observed relative nucleophilicities of these functionalities in the same MOF. When immobilized individually in monofunctional MOFs, the amine-functionalized linker was more reactive than the hydroxyl linker, indicating that the reactivity reversal observed in KSU-1 is due to the groups' mutual confinement within the MOF.