We have rationally designed a peptide that assembles into a redox-responsive, antimicrobial metallohydrogel. The resulting self-healing material can be rapidly reduced by ascorbate under physiological conditions and demonstrates a remarkable 160-fold change in hydrogel stiffness upon reduction. We provide a computational model of the hydrogel, explaining why position of nitrogen in non-natural amino acid pyridyl-alanine results in drastically different gelation properties of peptides with metal ions. Given its antimicrobial and rheological properties, the newly designed hydrogel can be used for removable wound dressing application, addressing a major unmet need in clinical care.

We have developed the stochastic microscopic-order-macroscopic-disorder (MOMD) approach for elucidating dynamic structures in the solid-state from H-2 NMR lineshapes. In MOMD, the probe experiences an effective/collective motional mode. The latter is described by a potential, u, which represents the local spatial-restrictions, a local-motional diffusion tensor, R, and key features of local geometry. Previously we applied MOMD to the well-structured core domain of the 3-fold-symmetric twisted polymorph of the A beta(40)-amyloid fibril. Here, we apply it to the N-terminal domain of this fibril. We find that the dynamic structures of the two domains are largely similar but differ in the magnitude and complexity of the key physical parameters. This interpretation differs from previous multisimple-mode (MSM) interpretations of the same experimental data. MSM used for the two domains different combinations of simple motional modes taken to be independent. For the core domain, MOMD and MSM disagree on the character of the dynamic structure. For the N-terminal domain, they even disagree on whether this chain segment is structurally ordered (MOMD finds that it is), and whether it undergoes a phase transition at 260 K where bulklike water located in the fibril matrix freezes (MOMD finds that it does not). These are major differences associated with an important system. While the MOMD description is a physically sound one, there are drawbacks in the MSM descriptions. The results obtained in this study promote our understanding of the dynamic structure of protein aggregates. Thus, they contribute to the effort to pharmacologically control neurodegenerative disorders believed to be caused by such aggregates.

During replication of herpesviruses, capsids escape from the nucleus into the cytoplasm by budding at the inner nuclear membrane. This unusual process is mediated by the viral nuclear egress complex (NEC) that deforms the membrane around the capsid by oligomerizing into a hexagonal, membrane-bound scaffold. Here, we found that highly basic membrane-proximal regions (MPRs) of the NEC alter lipid order by inserting into the lipid headgroups and promote negative Gaussian curvature. We also find that the electrostatic interactions between the MPRs and the membranes are essential for membrane deformation. One of the MPRs is phosphorylated by a viral kinase during infection, and the corresponding phosphomimicking mutations block capsid nuclear egress. We show that the same phosphomimicking mutations disrupt the NEC-membrane interactions and inhibit NEC-mediated budding in vitro, providing a biophysical explanation for the in vivo phenomenon. Our data suggest that the NEC generates negative membrane curvature by both lipid ordering and protein scaffolding and that phosphorylation acts as an off switch that inhibits the membrane-budding activity of the NEC to prevent capsid-less budding.IMPORTANCE Herpesviruses are large viruses that infect nearly all vertebrates and some invertebrates and cause lifelong infections in most of the world's population. During replication, herpesviruses export their capsids from the nucleus into the cytoplasm by an unusual mechanism in which the viral nuclear egress complex (NEC) deforms the nuclear membrane around the capsid. However, how membrane deformation is achieved is unclear. Here, we show that the NEC from herpes simplex virus 1, a prototypical herpesvirus, uses clusters of positive charges to bind membranes and order membrane lipids. Reducing the positive charge or introducing negative charges weakens the membrane deforming ability of the NEC. We propose that the virus employs electrostatics to deform nuclear membrane around the capsid and can control this process by changing the NEC charge through phosphorylation. Blocking NEC-membrane interactions could be exploited as a therapeutic strategy.

All radical S-adenosylmethionine (radical-SAM) enzymes, including the noncanonical radical-SAM enzyme diphthamide biosynthetic enzyme Dph1-Dph2, require at least one [4Fe-4S](Cys)(3) cluster for activity. It is well-known in the radical-SAM enzyme community that the [4Fe-4S](Cys)(3) cluster is extremely air-sensitive and requires strict anaerobic conditions to reconstitute activity in vitro. Thus, how such enzymes function in vivo in the presence of oxygen in aerobic organisms is an interesting question. Working on yeast Dph1-Dph2, we found that consistent with the known oxygen sensitivity, the [4Fe-4S] cluster is easily degraded into a [3Fe-4S] cluster. Remarkably, the small iron-containing protein Dph3 donates one Fe atom to convert the [3Fe-4S] cluster in Dph1-Dph2 to a functional [4Fe-4S] cluster during the radical-SAM enzyme catalytic cycle. This mechanism to maintain radical-SAM enzyme activity in aerobic environments is likely general, and Dph3-like proteins may exist to keep other radical-SAM enzymes functional in aerobic environments.

The development of antimicrobial peptides (AMPs) as potential therapeutics requires resolving the foundational principles behind their structure-activity relationships. The role of histidine residues within AMPs remains a mystery despite the fact that several potent peptides containing this amino acid are being considered for further clinical development. Gaduscidin-1 (Gad-1) is a potent AMP from Atlantic cod fish that has a total of five His residues. Herein, the role of His residues and metal-potentiated activity of Gad-1 was studied. The five His residues contribute to the broad-spectrum activity of Gad-1. We demonstrated that Gad-1 can coordinate two Cu2+ ions, one at the N-terminus and one at the C-terminus, where the C-terminal binding site is a novel Cu2+ binding motif. High affinity Cu2+ binding at both sites was observed using mass spectrometry and isothermal titration calorimetry. Electron paramagnetic resonance was used to determine the coordination environment of the Cu2+ ions. Cu2+ binding was shown to be responsible for an increase in antimicrobial activity and a new mode of action. Along with the traditional AMP mode of action of pore formation, Gad-1 in the presence of Cu2+ (per)oxidizes lipids. Importantly, His3, His11, His17, and His21 were found to be important to lipid (per) oxidation. This insight will help further understand the inclusion and role of His residues in AMPs, the role of the novel C-terminal binding site, and can contribute to the field of designing potent AMPs that bind metal ions to potentiate activity.

Light-induction of an anionic semiquinone (SQ) flavin radical in Drosophila cryptochrome (dCRY) alters the dCRY conformation to promote binding and degradation of the circadian clock protein Timeless (TIM). Specific peptide ligation with sortase A attaches a nitroxide spin-probe to the dCRY C-terminal tail (CTT) while avoiding deleterious side reactions. Pulse dipolar electron-spin resonance spectroscopy from the CTT nitroxide to the SQ shows that flavin photoreduction shifts the CTT similar to 1nm and increases its motion, without causing full displacement from the protein. dCRY engineered to form the neutral SQ serves as a dark-state proxy to reveal that the CTT remains docked when the flavin ring is reduced but uncharged. Substitutions of flavin-proximal His378 promote CTT undocking in the dark or diminish undocking in the light, consistent with molecular dynamics simulations and TIM degradation activity. The His378 variants inform on recognition motifs for dCRY cellular turnover and strategies for developing optogenetic tools.

Electrons added to TiO2 and other semiconductors often occupy trap states, whose reactivity can determine the catalytic and stoichiometric chemistry of the material. We previously showed that reduced aqueous colloidal TiO(2 )nanoparticles have two distinct classes of thermally equilibrated trapped electrons, termed Red/e(-) and Blue/e(-). Presented here are parallel optical and electron paramagnetic resonance (EPR) kinetic studies of the reactivity of these electrons with solution-based oxidants. Optical stopped-flow measurements monitoring the reactions of TiO2/e(-) with substoichiometric oxidants showed a surprising pattern: an initial fast (seconds) decrease in TiO2/e(-) absorbance followed by a secondary, slow (minutes) increase in the broad TiO2/e(-) optical feature. The analysis revealed that the fast decrease is due to the preferential oxidation of the Red/e(-) trap states and the slow increase results from the re-equilibration of electrons from Blue/e(-) to Red/e(-) states. This kinetic model was confirmed by freeze-quench EPR measurements. Quantitative analysis of the kinetic data demonstrated that Red/e(-) react similar to 5 times faster than Blue/e(-) with the nitroxyl radical oxidant 4-methoxy- 2,2,6,6-tetramethyl-1-piperidinyloxyl (4-MeO-TEMPO). Similar reactivity patterns were also observed in oxidations of TiO2/e(-) by O-2, which like 4-MeO-TEMPO is a proton-coupled electron transfer (PCET) oxidant, and by the pure electron transfer (ET) oxidant potassium triiodide (KI3). This suggests that the faster intrinsic reactivity of one trap state over another on the seconds-minutes time scale is likely a general feature of reduced TiO2 reactivity. This differential trap-state reactivity is likely to influence the performance of TiO2 in photochemical/electrochemical devices, and it suggests an opportunity for tuning catalysis.

In view of the great demand for using sustainable, renewable, and widely available resources to generate catalysts for advanced oxidation processes (AOPs), bio-originated waste materials show great promise as an alternative to state-of-the-art materials that suffer from high cost and scarcity. In this paper, we modified the ostrich bone waste (OBW) with hydrogen peroxide (HP) to produce OBW/HP with excellent chemical and mechanical strength that can in one sweep recover valuable heavy metal ions like Co from industrial wastewater and provide a sustainable catalyst-OBW/HP@CoNP- to activate peroxymonosulfate (PMS) for the degradation of dyes. After treatment of OBW with HP and Co ions, the morphologies of OBW/HP and OBW/HP@CoNP completely changed-porous, fine and flat sheets appeared, and well -dispersed, stable spherical Co nanoparticles of 50-70 nm were generated, respectively. PMSOBW/HP@CoNP system was able to remove several cationic and anionic dyes such as methylene blue (99.9 %), crystal violet (96.0 %), and congo red (76.0 %) within 120 min. As proof-of-concept applications, we prepared a membrane using a layer of OBW/HP@CoNP and a microreactor by encapsulating a water droplet within OBW/HP@CoNP shell, and found that the materials exhibit rapid kinetics to degrade MB within 30 s. In addition, the degradation of HEPES as a representative organic buffer was also studied for the first time at pH 7.4 and the mechanism of HEPES removal (98 %) was elucidated by LC-MS. Finally, although the preparation of OBW/HP@CoNP does not release toxic and synthetic compounds into the water, shows many advantages as catalyst and membrane- which promising its applications in industrial-scale including low-cost, sustainability, high chemical and mechanical stability, no metal leaching, and superior effectiveness in activating PMS, and ecyclability. (C) 2021 Institution of Chemical Engineers. Published by Elsevier B.V. All rights reserved.