Search Results (112)

A systematic review was conducted to assess the effects of microgravity and space radiation on astronauts’ oral health. This review aimed to determine if these conditions increase the risk of dental and periodontal diseases, identify pre-mission dental care strategies, and specify relevant dental emergencies for astronauts to manage during missions. Following PRISMA guidelines, the review was registered on PROSPERO (CRD42023472765). Databases including PubMed, Scopus, Web of Science, Cochrane Library, and OVID Medline were searched. Of the 13 studies identified, 7 were eligible for qualitative synthesis. The included studies revealed that space conditions compromise oral health. Findings indicate changes in saliva composition, with a significant decline in salivary lysozyme levels during missions lasting 28 to 84 days. Salivary IgA levels also increased before and peaked after flights (microgravity alters fluid shear and protein folding). Viral reactivation was a key finding, with latent viruses such as Epstein–Barr virus (EBV), cytomegalovirus (CMV), and varicella zoster virus (VZV) being reactivated during missions (immune suppression and gene expression shifts under spaceflight stress). Data from a study found that 50% of crew members shed viruses in their saliva or urine, and 38% tested positive for herpesviruses. The included studies also documented alterations in the oral microbiome, including increased gastrointestinal and decreased nasal microbial diversity. This suggests alterations in salivary biomarkers, viral shedding, and microbiome changes in astronauts during long-duration missions. These changes appear associated with immune dysregulation and stress, but causality remains uncertain due to observational designs, small heterogeneous samples, and confounding factors. Although current evidence is indicative rather than definitive, these findings highlight the need for preventive dental measures prior to missions and preparedness for managing oral emergencies in-flight. Future studies should address the mechanistic separation of microgravity and radiation effects, with implications for upcoming Moon and Mars missions. Full article

Adenosine triphosphate (ATP) is readily released into the extracellular space as an autocrine and paracrine purinergic signaling molecule. We originally reported a genetically encoded, fluorescent protein-based Förster Resonance Energy Transfer (FRET) biosensor that can detect micromolar levels of extracellular ATP. Through mutagenesis of the ATP binding site and optimization of cell-surface display, here we report the development of a second-generation biosensor called ECATS2 with greater than three-fold higher affinity for extracellular ATP. We found that the tether length between the FRET biosensor and the cell surface anchor is critical to optimization of its performance. Furthermore, we demonstrate that the improved sensor can detect extracellular ATP release upon hypoosmotic stress in cultured astrocytes. This new sensor contributes an improved tool for the ratiometric detection of extracellular ATP dynamics and purinergic signaling. Full article

The extraordinary diversity of protein sequences and structures gives rise to a vast protein functional universe with extensive biotechnological potential. Nevertheless, this universe remains largely unexplored, constrained by the limitations of natural evolution and conventional protein engineering. Substantial evidence further indicates that the known natural fold space is approaching saturation, with novel folds rarely emerging. AI-driven de novo protein design is overcoming these constraints by enabling the computational creation of proteins with customized folds and functions. This review systematically surveys the rapidly advancing field of AI-based de novo protein design, reviewing current methodologies and examining how cutting-edge computational frameworks accelerate discovery through three complementary vectors: (1) exploring novel folds and topologies; (2) designing functional sites de novo; (3) exploring sequence–structure–function landscapes. We highlight key applications across therapeutic, catalytic, and synthetic biology and discuss the persistent challenges. By fusing recent progress and the existing limitations, this review outlines how AI is not only accelerating the exploration of the protein functional universe but also fundamentally expanding the possibilities within protein engineering, paving the way for bespoke biomolecules with tailored functionalities. Full article

The cellular envelope of Gram-negative bacteria is a space where processes that are extremely important for the proper functioning of bacteria and determining their virulence take place. The extracytoplasmic protein quality control system, which includes chaperones, protein-folding catalysts, and proteases, is responsible for maintaining homeostasis in this cellular compartment. This system has been well studied in the model bacterium Escherichia coli, but little is known about its function in other bacteria. In bacteria evolutionarily distant from Enterobacteriaceae, the protein quality control system appears to function differently. For example, in the phylum Campylobacterota, a number of homologs of folding factors and proteases, whose functions are important for maintaining homeostasis in the periplasm of E. coli, have not been identified. Instead, there are quality control components that have no similar counterparts in the Enterobacteriaceae. In this review, we present the current state of knowledge on the extracytoplasmic protein quality control system in the model Campylobacterota, C. jejuni and H. pylori. Full article

The ZC4H2 gene is the site of congenital mutations linked to neurodevelopmental and musculoskeletal pathologies collectively termed ZARD (ZC4H2-Associated Rare Disorders). ZC4H2 consists of a coiled coil and a single novel zinc finger with four cysteines and two histidines, from which the protein obtains its name. Alpha Fold 3 confidently predicts a structure for the zinc finger but also for similarly sized random sequences, providing equivocal information on its folding status. We show using synthetic peptide fragments that the zinc finger of ZC4H2 is genuine and folds upon binding a zinc ion with picomolar affinity. NMR pH titration of histidines and UV–Vis of a cobalt complex of the peptide indicate its four cysteines coordinate zinc, while two histidines do not participate in binding. The experimental NMR structure of the zinc finger has a novel structural motif similar to RANBP2 zinc fingers, in which two orthogonal hairpins each contribute two cysteines to coordinate zinc. Most of the nine ZARD mutations that occur in the ZC4H2 zinc finger are likely to perturb this structure. While the ZC4H2 zinc finger shares the folding motif and cysteine-ligand spacing of the RANBP2 family, it is missing key substrate-binding residues. Unlike the NZF branch of the RANBP2 family, the ZC4H2 zinc finger does not bind ubiquitin. Since the ZC4H2 zinc finger occurs in a single copy, it is also unlikely to bind DNA. Based on sequence homology to the VAB-23 protein, the ZC4H2 zinc finger may bind RNA of a currently undetermined sequence or have alternative functions. Full article

Enzyme catalysis represents a promising approach for sustainable chemical synthesis, yet its industrial applications face limitations due to the inefficient regeneration and high cost of essential cofactors, such as adenosine-5′-triphosphate (ATP) and nicotinamide adenine dinucleotide phosphate (NADPH). While natural metabolic systems efficiently recycle cofactors through spatially organized enzymes, replicating this efficiency in vitro remains challenging. Here, we prepare a five-enzyme condensate system using liquid–liquid phase separation (LLPS) mediated by intrinsically disordered proteins (IDPs). By colocalizing a carboxylic acid reductase from Norcadia iowensis (NiCAR) with a reductive aminase from Aspergillus oryzae (AspRedAm) and three cofactor-regenerating enzymes, we generated a phase-separated catalytic condensate that enhanced ATP and NADPH recycling efficiency by 4.7-fold and 1.9-fold relative to free enzymes, respectively. Catalytic performance was correlated with the extent of phase separation, as confirmed by fluorescence microscopy, which revealed clear enrichment of ATP and NADPH within the condensates. This proximity effect enabled efficient cofactor turnover in the one-step reaction, achieving substrate conversion above 90% within 6 h and enhancing the space–time yield (STY) of the chiral imines 1.6-fold, with only one-fifth of the standard cofactor load. This approach creates a scalable and economic tool for performing multienzyme cascade reactions in vitro that are driven by the efficient recycling of multiple cofactors. Full article

The computational prediction of RNA three-dimensional (3D) structures remains a significant challenge, largely due to the limited understanding of RNA folding pathways. Although the scarcity of resolved native RNA structures has hindered the effectiveness of machine learning-based prediction methods, small, local structural motifs are both recurring and abundant in the available data. Precisely modeling these geometric motifs presents a promising approach to improving 3D structure prediction. In this paper, we introduce a novel mathematical model that represents RNA 3D structures as collections of interacting helices with concise geometric descriptions. By using a small set of parameters for each modeled helix, our method maps RNA strand segments onto helices within a 3D space, facilitating the effective assembly of large RNA structures. Preliminary tests on RNA sequences from the Protein Data Bank demonstrated the model’s potential in predicting key structural elements, including double helices, hairpin loops, and bulges. Full article

Direct methods based on iterative projection algorithms can determine protein crystal structures directly from X-ray diffraction data without prior structural information. However, traditional direct methods often converge to local minima during electron density iteration, leading to reconstruction failure. Here, we present an enhanced direct method incorporating genetic algorithms for electron density modification in real space. The method features customized selection, crossover, and mutation strategies; premature convergence prevention; and efficient message passing interface (MPI) parallelization. We systematically tested the method on 15 protein structures from different space groups with diffraction resolutions of 1.35∼2.5 Å. The test cases included high-solvent-content structures, high-resolution structures with medium solvent content, and structures with low solvent content and non-crystallographic symmetry (NCS). Results showed that the enhanced method significantly improved success rates from below 30% to nearly 100%, with average phase errors reduced below 40°. The reconstructed electron density maps were of sufficient quality for automated model building. This method provides an effective alternative for solving structures that are difficult to predict accurately by AlphaFold3 or challenging to solve by molecular replacement and experimental phasing methods. The implementation is available on Github. Full article

Water exists in the beginning and hydrates all matter. Life emerged in water, requiring three essential components in compartmentalized spaces: (1) universal energy sources driving biochemical reactions and processes, (2) molecules that store, encode, and transmit information, and (3) functional players carrying out biological activities and structural organization. Phosphorus has been selected to create adenosine triphosphate (ATP) as the universal energy currency, nucleic acids for genetic information storage and transmission, and phospholipids for cellular compartmentalization. Meanwhile, proteins composed of 20 α-amino acids have evolved into extremely diverse three-dimensional forms, including folded domains, intrinsically disordered regions (IDRs), and membrane-bound forms, to fulfill functional and structural roles. This review examines several unique findings: (1) insoluble proteins, including membrane proteins, can become solubilized in unsalted water, while folded cytosolic proteins can acquire membrane-inserting capacity; (2) Hofmeister salts affect protein stability by targeting hydration; (3) ATP biphasically modulates liquid–liquid phase separation (LLPS) of IDRs; (4) ATP antagonizes crowding-induced protein destabilization; and (5) ATP and triphosphates have the highest efficiency in inducing protein folding. These findings imply the following: (1) hydration might be encoded in protein sequences, central to manifestation and modulation of protein structures, dynamics, and functionalities; (2) phosphate anions have a unique capacity in enhancing μs-ms protein dynamics, likely through ionic state exchanges in the hydration shell, underpinning ATP, polyphosphate, and nucleic acids as molecular chaperones for protein folding; and (3) ATP, by linking triphosphate with adenosine, has acquired the capacity to spacetime-specifically release energy and modulate protein hydration, thus possessing myriad energy-dependent and -independent functions. In light of the success of AlphaFolds in accurately predicting protein structures by neural networks that store information as distributed patterns across nodes, a fundamental question arises: Could cellular networks also handle information similarly but with more intricate coding, diverse topological architectures, and spacetime-specific ATP energy supply in membrane-compartmentalized aqueous environments? Full article

In recent years, the increasing number of long-duration space missions has prompted the scientific community to undertake a more comprehensive examination of the impact of microgravity on the human body during spaceflight. This review aims to assess the current knowledge regarding the consequences of exposure to an extreme environment, like microgravity, on the human body, focusing on the role of heat-shock proteins (HSPs). Previous studies have demonstrated that long-term exposure to microgravity during spaceflight can cause various changes in the human body, such as muscle atrophy, changes in muscle fiber composition, cardiovascular function, bone density, and even immune system functions. It has been postulated that heat-shock proteins (HSPs) may play a role in mitigating the harmful effects of microgravity-induced stress. According to past studies, heat-shock proteins (HSPs) are upregulated under simulated microgravity conditions. This upregulation assists in the maintenance of the proper folding and function of other proteins during stressful conditions, thereby safeguarding the physiological systems of organisms from the detrimental effects of microgravity. HSPs could also be used as biomarkers to assess the level of cellular stress in tissues and cells exposed to microgravity. Therefore, modulation of HSPs by drugs and genetic or environmental techniques could prove to be a potential therapeutic strategy to reduce the negative physiological consequences of long-duration spaceflight in astronauts. Full article

Background: Neutrophils eject their DNA strings and cellular proteins into the extracellular space upon treatment with various stimulants. In the present study, we examined the effects of four major oxidized cholesterol metabolites on DNA release from granulocytes. Methods and Results: When oxysterols were added to HL-60-derived granulocytes stimulated with phorbol 12-myristate 13-acetate (PMA), they suppressed the release of DNA and myeloperoxidase from the cells. Among the four oxysterols tested, 7-ketocholesterol was the most effective. Addition of the same concentration of 7-ketocholesterol did not induce any cytotoxic effects, as evaluated based on the release of lactate dehydrogenase and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazoliumbromide (MTT) assays. DNA release from human peripheral blood neutrophils after PMA stimulation was also suppressed by 7-ketocholesterol. Liquid chromatography with tandem mass spectrometry (LC-MS/MS) analysis was used to quantify sterol content in the cells. The addition of oxysterols increased the cellular content of the corresponding compounds by more than 10-fold compared to those at baseline. Treatment of HL-60-derived granulocytes with methyl-β-cyclodextrin that removes sterol compounds from the membranes increased DNA release from the cells in a dose-dependent manner. Conclusions: These results suggest that oxysterols have suppressive effects on DNA release from granulocytes stimulated with PMA. Full article

Background: Ricin, a toxin extracted from the seeds of Ricinus communis, is classified as a ribosome-inactivating protein. The A-subunit of ricin shows RNA N-glycosidase activity that cleaves ribosomal RNA (rRNA) and exhibits toxicity by inhibiting protein synthesis and inducing vascular leak syndrome. Methods: In this study, we created a truncated version of the previously developed R51 ricin vaccine (RTA 1-194 D75C Y80C) through in silico analysis. Results: The resulting R51-3 vaccine showed a more-than-six-fold increase in soluble protein expression when compared to R51, with over 85% solubility. In a pilot toxicity test, no toxicity was observed in hematological and biochemical parameters in BALB/c mice and New Zealand white rabbits following five repeated administrations of R51-3. Furthermore, R51-3 successfully protected mice and rabbits from a 20 × LD₅₀ ricin challenge after three intramuscular injections spaced 2 weeks apart. Similarly, monkeys that received three injections of R51-3 survived a 60 µg/kg ricin challenge. Conclusions: These findings support R51-3 as a promising candidate antigen for ricin vaccine development. Full article

At every point $p$ on a smooth $n$-manifold M there exist $\left(n+1\right)$ skew-symmetric tensor spaces spanning differential $r$-forms $\omega$ with $r=0,1,\cdots ,n$. Because $d\circ d$ is always zero where $d$ is the exterior differential, it follows that every exact $r$-form (i.e., $\omega =d\lambda$ where $\lambda$ is an $\left(r-1\right)$-form) is closed (i.e., $d\omega =0$) but not every closed $r$-form is exact. This implies the existence of a third type of differential $r$-form that is closed but not exact. Such forms are called harmonic forms. Every smooth $n$-manifold has an underlying topological structure. Many different possible topological structures exist. What distinguishes one topological structure from another is the number of holes of various dimensions it possesses. De Rham’s theory of differential forms relates the presence of $r$-dimensional holes in the underlying topology of a smooth $n$-manifold $M$ to the presence of harmonic $r$-form fields on the smooth manifold. A large amount of theory is required to understand de Rham’s theorem. In this paper we summarize the differential geometry that links holes in the underlying topology of a smooth manifold with harmonic fields on the manifold. We explore the application of de Rham’s theory to (i) visual, (ii) mechanical, (iii) electrical and (iv) fluid flow systems. In particular, we consider harmonic flow fields in the intracellular aqueous solution of biological cells and we propose, on mathematical grounds, a possible role of harmonic flow fields in the folding of protein polypeptide chains. Full article

Reliable and accurate methods of estimating the accuracy of predicted protein models are vital to understanding their respective utility. Discerning how the quaternary structure conforms can significantly improve our collective understanding of cell biology, systems biology, disease formation, and disease treatment. Accurately determining the quality of multimeric protein models is still computationally challenging, as the space of possible conformations is significantly larger when proteins form in complex with one another. Here, we present EGG (energy and graph-based architectures) to assess the accuracy of predicted multimeric protein models. We implemented message-passing and transformer layers to infer the overall fold and interface accuracy scores of predicted multimeric protein models. When evaluated with CASP15 targets, our methods achieved promising results against single model predictors: fourth and third place for determining the highest-quality model when estimating overall fold accuracy and overall interface accuracy, respectively, and first place for determining the top three highest quality models when estimating both overall fold accuracy and overall interface accuracy. Full article

Bacitracin Methylene Disalicylate (BMD), as a feed additive to poultry diets, enhances digestion, prevents Salmonella enteritidis (SE) colonization, and treats current infections. The objective of this study was to utilize a quantitative proteomic approach to determine the effect of BMD feed additive on broiler chickens challenged with SE in the spleen proteome. At 1 d of age, chicks were randomly allocated into four groups: control with and without SE challenge (CON, n = 60; CON-SE, n = 60), BMD with and without SE challenge (BMD, n = 60; BMD-SE, n = 60). Birds in the CON-SE and BMD-SE treatment were administered SE inoculum by oral gavage. On day three and day seven post-gavage, the spleen was collected aseptically from birds in each treatment group (CON, n = 4/day; CON-SE, n = 4/day; BMD, n = 4/day; BMD-SE, n = 4/day). Proteomic analysis by liquid chromatography-tandem mass spectrometry (LC-MS/MS) showed an increased abundance of 115 proteins and decreased of 77 due to the BMD. Proteins that decreased in abundance were enriched for fibrinogen complex and extracellular space, whereas proteins that increased in abundance were enriched for proteasome-mediated ubiquitin-dependent protein catabolic process and mitochondrion. Analysis of the interaction between BMD and the Salmonella challenge found 230 differentially abundant proteins including proteins associated with RNA binding, spliceosome, protein transport, and cell adhesion among the upregulated proteins, and those associated with protein folding, carbon metabolism, biosynthesis of nucleotide sugars, response to oxidative stress, positive regulation of NIK/NF-kappaB signaling, and inflammatory response among the downregulated proteins. The impact of BMD treatment on spleen proteome indicates an anti-apoptotic effect. BMD also modified the response of the spleen to the SE challenge with a marked decrease in proteins that prompt cytokine synthesis and an increase in proteins involved in the selective removal of unfolded proteins. Full article