Urate oxidase (Uox) has great potential in biotechnology. However, the exploration of Uox with superior stability and its simple preparation with high yield remains challenging. By fusing human ferritin with the Uox from the extremophile Deinococcus radiodurans, a novel 24-polymeric Uox (DrUox-F) with excellent properties was achieved. The DrUox-F with 98 % purity was obtained through low-speed centrifugation with the yield of 890 mg/L, representing the highest yield currently reported. Remarkably, DrUox-F preserved 100 % activity after 35 days at pH 10, which was significantly superior to DrUox. Furthermore, no significant activity loss was observed following 318 days of storage at 4 °C. The lyophilized DrUox-F power retained over 90 % of original activity after 280 days of storage at room temperature. Notably, the half-life was extended approximately 40-fold (from 41 h to 1880 h) through the addition of mannitol as an exogenous stabilizer. Molecular docking results revealed that mannitol could specifically bind to multiple regions on DrUox-F, with hydrogen bonds as the dominant interactions. Thus, DrUox-F, with superior cost-effectiveness, high efficiency, and enhanced stability, provides a breakthrough solution for addressing the critical limitations of Uox.

Carbonyl reductases (CRs) are important enzymes with a broad spectrum of industrial applications. However, the CRs reported for use in the asymmetric synthesis of chiral aryl β-hydroxy α-amino acids are exclusively specific to NADPH, posing challenges for cost-effective NADH regeneration. In this study, we focused on the carbonyl reductase (M30) from Exiguobacterium sp. F42 and successfully altered its cofactor preference to favor the more economical NADH using a computer-aided design approach. Through protein engineering, we developed an enhanced mutant M36, S10A/Y15R/E16A/K19L/A32D/R33I, which increased the cofactor specificity of M30 by over 1000-fold. This best combinatorial mutant enabled the highly efficient synthesis of the chloramphenicol intermediate (2S,3R)-5b, achieving 99 % conversion from 50 g/L of substrate 5a with excellent stereoselectivity (ee > 99.9 %, de > 99.9 %), hereby significantly enhancing its industrial application potential. Molecular dynamics (MD) simulations provided atomic-level insights into the mechanistic basis for the altered cofactor affinity and revealed the individual contribution of key mutations. This work establishes an efficient approach for engineering cofactor specificity in CRs and underscores the potential of cavity engineering in advancing biocatalytic applications.

AbstractSilent information regulator 2 (Sir2) proteins typically catalyze NAD+-dependent protein deacetylation. The recently identified bacterial Sir2 domain-containing protein, defense-associated sirtuin 2 (DSR2), recognizes the phage tail tube and depletes NAD+ to abort phage propagation, which is counteracted by the phage-encoded DSR anti-defense 1 (DSAD1), but their molecular mechanisms remain unclear. Here, we determine cryo-EM structures of inactive DSR2 in its apo form, DSR2–DSAD1 and DSR2–DSAD1–NAD+, as well as active DSR2–tube and DSR2–tube–NAD+ complexes. DSR2 forms a tetramer with its C-terminal sensor domains (CTDs) in two distinct conformations: CTDclosed or CTDopen. Monomeric, rather than oligomeric, tail tube proteins preferentially bind to CTDclosed and activate Sir2 for NAD+ hydrolysis. DSAD1 binding to CTDopen allosterically inhibits tube binding and tube-mediated DSR2 activation. Our findings provide mechanistic insight into DSR2 assembly, tube-mediated DSR2 activation, and DSAD1-mediated inhibition and NAD+ substrate catalysis in bacterial DSR2 anti-phage defense systems.

Neuronal activation is required for the formation of drug-associated memory, which is critical for the development, persistence, and relapse of drug addiction. Nevertheless, the metabolic mechanisms underlying energy production for neuronal activation remain poorly understood. In the study, a large-scale proteomics analysis of lysine crotonylation (Kcr), a type of protein posttranslational modification (PTM), reveals that cocaine promoted protein Kcr in the hippocampal dorsal dentate gyrus (dDG). We find that Kcr is predominantly discovered in a few enzymes critical for mitochondrial energy metabolism; in particular, pyruvate dehydrogenase (PDH) complex E1 subunit α (PDHA1) is crotonylated at the lysine 39 (K39) residue through P300 catalysis. Crotonylated PDHA1 promotes pyruvate metabolism by activating PDH to increase ATP production, thus providing energy for hippocampal neuronal activation and promoting cocaine-associated memory recall. Our findings identify Kcr of PDHA1 as a PTM that promotes pyruvate metabolism to enhance neuronal activity for cocaine-associated memory.

The Bin/Amphiphysin/Rvs (BAR) domain protein FAM92A1 is a multifunctional protein engaged in regulating mitochondrial ultrastructure and ciliogenesis, but its physiological role in the brain remains unclear. Here, we show that FAM92A1 is expressed in neurons starting from embryonic development. FAM92A1 knockout in mice results in altered brain morphology and age-associated cognitive deficits, potentially due to neuronal degeneration and disrupted synaptic plasticity. Specifically, FAM92A1 deficiency impairs diverse neuronal membrane morphology, including the mitochondrial inner membrane, myelin sheath, and synapses, indicating its roles in membrane remodeling and maintenance. By determining the crystal structure of the FAM92A1 BAR domain, combined with atomistic molecular dynamics simulations, we uncover that FAM92A1 interacts with phosphoinositide- and cardiolipin-containing membranes to induce lipid-clustering and membrane curvature. Altogether, these findings reveal the physiological role of FAM92A1 in the brain, highlighting its impact on synaptic plasticity and neural function through the regulation of membrane remodeling and endocytic processes.

Electro-osmosis has been well recognized as a technique for the remediation of petroleum-contaminated soil, however seasonally freezing and thawing adds the complexity of petroleum mobility in cold regions. To investigate the influence of freeze-thaw on the electroosmotic removal of petroleum and explore the enhancement of freeze-thaw on the electroosmotic remediation efficiency in remediating the petroleum-contaminated soils, a set of laboratory tests were performed in three types of treatment modes, freeze-thaw (FT), electro-osmosis (EO) and freeze-thaw combined electro-osmosis (FE). The petroleum redistributions as well as the moisture content changes after the treatments were evaluated and compared. The petroleum removal rates of the three treatments were analyzed, and the underlying mechanisms were elaborated. The results indicated that the overall efficiency of the treatment mode regarding petroleum removal from soil followed the order of FE > EO > FT, corresponding to 54%, 36% and 21% in maximum, respectively. A considerable amount of water solution with surfactant was driven into contaminated soil during FT process, but the petroleum mobilization primarily occurred inside of the specimen. A higher remediation efficiency was yield in EO mode, but the induced dehydration and cracks leaded to the dramatical depression in the efficiency in further process. It is proposed that the petroleum removal is closely related to the flow of water solution with surfactant that is favorable to the solubility and mobilization of the petroleum in soil. Thus, the water migration induced by freeze-thaw cycles substantially improved the efficiency of the electroosmotic remediation in FE mode that gave the best performance for the remediation of the petroleum-contaminated soil.

Supplementary Figures 1-8 Supplementary Table 1

Movie S. AMFR RING (cyan) and B7-H4 dimer complex in 120 ns MD simulations (run 1). Simulations were performed to verify the stability of the complex conformer illustrated in Fig. 4E. B7-H4 Ig-like V-type 1 and Ig-like V-type 2 domains are in pale green and green. Ubiquitination sites (K138 and K146) are highlighted in blue spheres. Asparagines are shown in orange sticks. Interacting residues between the two proteins are represented as CPK and colored by atom types.

Warm frozen soil is currently the subject of research by many researchers in cold regions. Its highly unstable mechanical properties, particularly its dynamic properties, play an essential role in the instability of frozen soil foundations. The deformation of warm frozen soils and pore-water pressure are related, the confined compression tests under dynamic load were performed to clarify the relationship between them. And the variation law of pore-water pressure and deformation in frozen soils under the effect of four factors: temperature, dynamic stress amplitude, dry density and frequency were discussed, and the mechanism of pore-water pressure and deformation is also elaborated. The results show that the pore-water pressure of frozen soils varies greatly during the test, with a general three-stage pattern of increasing-dissipating-increasing. And the deformation of frozen soil has a significant regularity, and the overall variation trend is rapid increase followed by dynamic stability or slow growth. Meanwhile, creep and consolidation deformation are important components of frozen soil deformation, and the deformation generated by both in the test basically accounts for about 50% or more of the total deformation, and in some cases even up to about 64%. In addition, it is indicated that there is a positive linear relationship between the deformation of the specimen and the pore-water pressure. In conclusion, it can be helpful to carry out work in cold regions safely by studying the variation of pore-water pressure and deformation characteristics of warm frozen soils under dynamic loading.

Objective: Contextual drug-associated memory precipitates craving and relapse in substance users, and the risk of relapse is a major challenge in the treatment of substance use disorders. Thus, understanding the neurobiological underpinnings of how this association memory is formed and maintained will inform future advances in the treatment of drug addiction. Brain endocannabinoids (eCBs) signalling has been associated with drug-induced neuroadaptations, but the role of lipases that mediate small lipid ligand biosynthesis and metabolism in regulating drug-associated memory has not been examined. Here, we explored how manipulation of the lipase fatty acid amide hydrolase (FAAH), which is involved in mediating the level of the lipid ligand anandamide (AEA), affects cocaine-associated memory formation.Methods: We applied behavioural, pharmacological and biochemical methods to detect cocaine-associated memory formation, eCBs in the dorsal dentate gyrus (dDG), and the activity of related enzymes. We further examined the roles of abnormal FAAH activity and AEA-CB1R signalling in the regulation of cocaine-associated memory formation and granule neuron dendritic structure alterations in the dDG through Western blotting, electron microscopy and immunofluorescence.Results: In the present study, we found that cocaine induced a decrease in the level of FAAH in the dDG and increased the level of AEA. A high level of AEA activated cannabinoid type 1 receptors (CB1Rs) and further triggered CB1R signalling activation and granule neuron dendritic remodelling, and these effects were reversed by blockade of CB1Rs in the brain. Furthermore, inhibition of FAAH in the dDG markedly increased AEA levels and promoted cocaine-associated memory formation through CB1R signalling activation. Conclusions: Together, our findings demonstrate that the lipase FAAH influences CB1R signalling activation and granule neuron dendritic structure alteration in the dDG by regulating AEA levels and that AEA and AEA metabolism play a key role in cocaine-associated memory for-mation. Manipulation of AEA production may serve as a potential therapeutic strategy for drug addiction and relapse prevention.O 2022 The Author(s). Published by Elsevier GmbH. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).

The systematic design of functional peptides has technological and therapeutic applications. However, there is a need for pattern-based search engines that help locate desired functional motifs in primary sequences regardless of their evolutionary conservation. Existing databases such as The Protein Secondary Structure database (PSS) no longer serves the community, while the Dictionary of Protein Secondary Structure (DSSP) annotates the secondary structures when tertiary structures of proteins are provided. Here, we extract 1.7 million helices from the PDB and compile them into a database (Therapeutic Peptide Design database; TP-DB) that allows queries of compounded patterns to facilitate the identification of sequence motifs of helical structures. We show how TP-DB helps us identify a known purification-tag-specific antibody that can be repurposed into a diagnostic kit for Helicobacter pylori. We also show how the database can be used to design a new antimicrobial peptide that shows better Candida albicans clearance and lower hemolysis than its template homologs. Finally, we demonstrate how TP-DB can suggest point mutations in helical peptide blockers to prevent a targeted tumorigenic protein-protein interaction. TP-DB is made available at http://dyn.life.nthu.edu.tw/design/ .

Sedimentary nitrogen isotope (δ15Nsed) in Santa Barbara Basin (SBB) has been mostly interpreted as the record of the eastern tropical North Pacific (ETNP) intermediate water denitrification process. Nevertheless, debate remains regarding sources and control mechanisms of δ15Nsed signal in SBB. Multi-proxy analyses including δ15Nsed, total organic carbon (TOC), total nitrogen (TN), C/N ratio, and marine biomarkers were performed on a 46-cm sediment core (SBB-190629) collected from SBB in 2019. The core was dated with varve counting and 210Pb dating method, showing a depositional history of 1938–2019 CE with a sedimentation rate of 0.564 cm/year. The findings show that the δ15Nsed record (at ~0.25-year resolution) ranges from 6.24‰ to 7.43‰, which was affected by both local and remote forcing. The long-term variations of the SBB δ15Nsed signature show a general decreasing trend from 1940 to the late 1980s, low values during 1980~2000, and an increase afterward, which is thought to reflect changes in ETNP denitrification induced by the strength of tropical trade winds. Our results also reveal a series of abrupt annual to multiannual changes, superimposed on the long-term variation mentioned above. The SBB local δ15N signal (Δδ15NSBB) is accessed by using the deviation from the mean δ15N (Δδ15N) of SBB-190629 to subtract the Δδ15N of the ETNP. The Δδ15NSBB record compares well with redox-sensitive proxies (Re/Mo ratio and C29 stanol/stenol ratio) from the SBB bottom water and with the OCmarine content calculated based on the C/N ratio mixing model, indicating that the Δδ15NSBB is mainly controlled by bottom water denitrification, which was induced by the change of upwelling intensity and marine productivity. Since various climatic factors (e.g., El Niño–Southern Oscillation (ENSO), Pacific Decadal Oscillation (PDO), and North Pacific Gyre Oscillation (NPGO)) have different impacts on the upwelling intensity (hence the marine productivity and denitrification) in SBB on different timescales, the influence of combined climatic factors on SBB denitrification is time dependent.

Alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptors (AMPARs) are key regulators during the process of synaptic plasticity in major depression disorder (MDD). Synapse differentiation-induced gene 1 (SynDIG1) functions as an atypical AMPAR auxiliary subunit and regulates synaptic AMPAR content; however, the role of SynDIG1 in MDD remains elusive. In this study, we found that the SynDIG1 expression was significantly increased in the neurons of the nucleus accumbens (NAc) of male mice after chronic social defeat stress (CSDS). CSDS enhanced SynDIG1-GluA2 binding and promoted the surface expression of AMPAR subunit GluA2 in the NAc. Knockdown of SynDIG1 decreased the surface expression of GluA2 and reversed the alteration of dendrite spines in the neurons, eventually alleviating the depressive-like behaviors of the stressed mice. Moreover, intra-NAc injection of IP12, a specific peptide to disrupt the interaction of SynDIG1 with GluA2, rescued depressive-like behaviors. Collectively, SynDIG1 regulates the surface expression of GluA2 and dendritic remodeling in the NAc of male mice under CSDS, thus mediating the depressive-like behaviors.

In this study, closed-field unbalanced magnetron sputtering was employed to deposit a diamond-like carbon (DLC) film with Cr as the middle layer on three elastomeric substrates, namely nitrile rubber (NBR), fluorine rubber (FKM), and ethylene propylene diene monomer (EPDM) rubber, respectively. In addition, effects of these rubber substrates on the structure and properties of DLC films were investigated. Results revealed that after the bending test, several new cracks are produced on the DLC-coated EPDM rubber and NBR rubber, while debonding is not observed, indicative of the good flexibility and adhesion of DLC films on EPDM rubber and NBR rubber. Furthermore, tribological properties of NBR rubber and EPDM rubber were significantly improved by the DLC film, and almost no film damage was observed on the wear track, while the DLC coated-FKM exhibited severe wear due to the gradual peeling of the coating. Film shedding was caused by the extremely smooth FKM substrate, which led to low film adhesion. Different tribological properties of the coated rubbers were closely related to the difference in surface roughness and mechanical properties of the rubber substrates.

Abstract Summary ProDy, an integrated application programming interface developed for modelling and analysing protein dynamics, has significantly evolved in recent years in response to the growing data and needs of the computational biology community. We present major developments that led to ProDy 2.0: (i) improved interfacing with databases and parsing new file formats, (ii) SignDy for signature dynamics of protein families, (iii) CryoDy for collective dynamics of supramolecular systems using cryo-EM density maps and (iv) essential site scanning analysis for identifying sites essential to modulating global dynamics. Availability and implementation ProDy is open-source and freely available under MIT License from https://github.com/prody/ProDy. Supplementary information Supplementary data are available at Bioinformatics online.

EDITORIAL article Front. Mol. Biosci., 21 April 2021 | https://doi.org/10.3389/fmolb.2021.681364

Multiple sclerosis (MS) is the most common chronic inflammatory demyelinating disease of the central nervous system. While the drugs currently available for MS provide symptomatic benefit, there is no curative treatment. The emergence of large-scale multiomics data and network theory provide new opportunities for drug discovery in MS, as these are promising strategies for developing novel drugs. In this study, we proposed a computational framework that combined biomolecular network modeling and structural dynamics analysis to facilitate the discovery of new drugs with potential activity in MS. First, we developed a new shortest path-based algorithm that prioritized differentially expressed genes using a newly topological and functional exploration of protein-protein interaction network. Then, pathway enrichment analysis and an assessment of target druggability suggested that TNF-α-induced protein 3 (TNFAIP3), which is involved in NF-κ B signaling, could be a potential therapeutic target for MS. Finally, druggability simulations and mutation enrichment analysis of the TNFAIP3 dimer presented two druggable sites. Follow-up pharmacophore model-based virtual screening of the two sites yielded 30 hit compounds with low energy scores. In summary, this novel method based on analyzing "omics data" and performing druggability simulations, is a systematic approach that unravels disease mechanisms and links them to the chemical space to develop treatments and can be applied to other complex diseases.

Protein–ligand interaction analysis is important for drug discovery and rational protein design. The existing online tools adopt only a single conformation of the complex structure for calculating and displaying the interactions, whereas both protein residues and ligand molecules are flexible to some extent. The interactions evolved with time in the trajectories are of greater interest. MolADI is a user-friendly online tool which analyzes the protein–ligand interactions in detail for either a single structure or a trajectory. Interactions can be viewed easily with both 2D graphs and 3D representations. MolADI is available as a web application.

Hydrogen polysulfide (H 2 S n , n >1), as a direct oxidizing form of hydrogen sulfide, is closely associated with intestinal diseases such as ulcerative colitis (UC).