Prokaryotic Research Articles

Pola Bakteri Aerob dan Uji Kepekaan Antibiotik pada Urine Pasien dengan Kateter Uretra di RSU Gmim Pancaran Kasih Manado

Bacteria are prokaryotic organisms, many of which are part of the normal flora in the human body without causing infection. The infection pattern caused by this aerobic bacteria can very depending on factors such as the duration of catheter use, the patient’s health condition, and hygiene levels. Catheter-Associated Urinary Tract Infections (CAUTI) is one of the most common infections in the ICU and contribute to increased mortality and morbidity rates. Objective: To determine the aerobic bacterial patterns in urethral catheters and their antibiotic susceptibility at Pancaran Kasih General Hospital, Manado. Method: This study used a descriptive observational design with a cross-sectional approach to analyze the aerobic bacterial patterns and antibiotic susceptibility in the urine of patients with urethral catheters at GMIM Pancaran Kasih General Hospital, Manado. Results: Samples were taken from patients using urethral catheters, and 9 Gram-positive and 5 Gram-negative bacteria were identified, with Staphylococcus sp. being the most frequently found bacterium. Antibiotic susceptibility testing showed that Staphylococcus sp., Staphylococcus aureus, and Bacillus sp. were highly sensitive to two antibiotics: doxycycline and meropenem. Conversely, these bacteria showed resistance to several antibiotics, including chloramphenicol. Conclusion: Urethral catheter use revealed Staphylococcus sp. as the most frequently found bacterium. Antibiotic susceptibility testing showed that Staphylococcus sp., Staphylococcus aureus, and Bacillus sp. were sensitive to doxycycline and meropenem.

Unravelling Faecal Microbiota Variations in Equine Atypical Myopathy: Correlation with Blood Markers and Contribution of Microbiome

Hypoglycin A and methylenecyclopropylglycine are protoxins responsible for atypical myopathy in equids. These protoxins are converted into toxins that inhibit fatty acid β-oxidation, leading to blood accumulation of acylcarnitines and toxin conjugates, such as methylenecyclopropylacetyl-carnitine. The enzymes involved in this activation are also present in some prokaryotic cells, raising questions about the potential role of intestinal microbiota in the development of intoxication. Differences have been noted between the faecal microbiota of cograzers and atypical myopathy-affected horses. However, recent blood acylcarnitines profiling revealed subclinical cases among cograzers, challenging their status as a control group. This study investigates the faecal microbiota of horses clinically affected by atypical myopathy, their cograzers, and a control group of toxin-free horses while analysing correlations between microbiota composition and blood parameters. Faecal samples were analysed using 16S amplicon sequencing, revealing significant differences in α-diversity, evenness, and β-diversity. Notable differences were found between several genera, especially Clostridia_ge, Bacteria_ge, Firmicutes_ge, Fibrobacter, and NK4A214_group. Blood levels of methylenecyclopropylacetyl-carnitine and C14:1 correlated with variations in faecal microbial composition. The theoretical presence of enzymes in bacterial populations was also investigated. These results underscore the critical need to investigate the potential role of intestinal microbiota in this poisoning and may provide insights for developing prevention and treatment strategies.

A framework for understanding and investigating polyphosphate-protein interactions.

Many prokaryotic and eukaryotic cells store inorganic phosphate in the form of polymers called polyphosphate (polyP). There has been an explosion of interest in polyP over the past decade, in part due to newly suggested roles related to diverse aspects of human health. The physical interaction of polyP chains with specific proteins has been proposed to regulate cellular homeostasis and modulate signaling pathways in response to environmental changes. Recently, several studies have challenged existing models for how polyP interacts with its protein targets, while identifying new motifs that are capable of binding to polyP. In this review, we summarize these findings, delineate the functional implications for polyP-protein interactions at the molecular level, and define open questions that should be addressed to propel the field forward.

Novel Approaches to Decomposing Hydrocarbon Pollutants from the Environment

In the modern era, petrochemical industries' production of hydrocarbon pollution is a significant environmental problem that causes biodiversity loss. Alkanes constitute a substantial proportion of crude oil, and refined fuels are found in small amounts in various uncontaminated environments. They are prevalent in underground fossil fuel reserves and shallow subsurface habitats polluted with hydrocarbons, such as aquifers. Using microorganisms to break down alkane hydrocarbon pollutants in environmental areas has great potential. Considerable advancements have been achieved in identifying microorganisms and metabolic processes responsible for the breakdown of alkanes in both oxygen-free and oxygen-rich conditions in the last two decades. A wide range of prokaryotic and eukaryotic organisms have been identified and observed to possess the ability to utilize various carbon and energy sources as substrates. Bioremediation is essential for environmental safety and management; various methods have been established for petroleum hydrocarbon bioremediation. Numerous microbial species have been employed to investigate the bioremediation of petroleum hydrocarbons, highlighting the crucial functions of varied microbial communities. Phytoremediation is an environmentally sustainable method that may effectively rehabilitate heavy metal- contaminated soil cost-efficiently. This manuscript provides an overview of prevalent alkane hydrocarbon pollutants, microorganisms capable of degrading hydrocarbons, key pathways and enzymes involved in hydrocarbon degradation, factors influencing hydrocarbon degradation, and various strategies employed to harness the degrading capabilities of microbes for remedial purposes.

The Dps Protein Protects Escherichia coli DNA in the Form of the Trimer.

The Dps protein is the major DNA-binding protein of prokaryotes, which protects DNA during starvation by forming a crystalline complex. The structure of such an intracellular DNA-Dps complex is still unknown. However, the phenomenon of a decrease in the size of the Dps protein from 90 Å to 69-75 Å during the formation of a complex with DNA has been repeatedly observed, and no explanation has been given. In this work, we show that during the formation of intracellular DNA-Dps crystals, the protein transitions to another oligomeric form: from a dodecameric (of 12 monomers), which has an almost spherical shape with a diameter of 90 Å, to a trimeric (of three monomers), which has a shape close to a torus-like structure with a diameter of 70 Å and a height of 40 Å. The trimer model was obtained through the molecular dynamic modeling of the interaction of the three monomers of the Dps protein. Placement of the obtained trimer in the electron density of in vitro DNA-Dps crystal allowed for the determination of the lattice parameters of the studied crystal. This crystal model was in good agreement with the SAXS data obtained from intracellular crystals of 2-day-old Escherichia coli cells. The final crystal structure contains a DNA molecule in the through channel of the crystal structure between the Dps trimers. It was discussed that the mechanism of protein transition from one oligomeric form to another in the cell cytoplasm could be regulated by intracellular metabolites and is a simple and flexible mechanism of prokaryotic cell transition from one metabolic state to another.

Directed evolution of an orthogonal transcription engine for programmable gene expression in eukaryotes.

T7 RNA polymerase (RNAP) has enabled orthogonal control of gene expression and recombinant protein production across diverse prokaryotic host chassis organisms for decades. However, the absence of 5' methyl guanosine caps on T7 RNAP-derived transcripts has severely limited its utility and widespread adoption in eukaryotic systems. To address this shortcoming, we evolved a fusion enzyme combining T7 RNAP with the single subunit capping enzyme from African swine fever virus using Saccharomyces cerevisiae. We isolated highly active variants of this fusion enzyme, which exhibited roughly two orders of magnitude higher protein expression compared to the wild-type enzyme. We demonstrate the programmable control of gene expression using T7 RNAP-based genetic circuits in yeast and validate enhanced performance of these engineered variants in mammalian cells. This study presents a robust, orthogonal gene regulatory system applicable across diverse eukaryotic hosts, enhancing the versatility and efficiency of synthetic biology applications.

THE EUKARYOTE ENDOSYMBIOTIC ORIGIN: A CONSTRUCTAL THEORY-BASED EXPLANATION

The origin of eukaryotes is understood as one of the most important issues in the history of living beings. There are several points of view to seek a broad understanding of the eukaryote origins, which encompass paleontological data, energetics, eukaryote-particular characteristics origins or the connections among dissimilar eukaryotic groups. Many proposals of endosymbiotic theory have been presented to explain the origin of eukaryotes and their mitochondria. Only recently, energy and the energetic constraints started to be considered by endosymbiotic theory to understand the contribution of prokaryotic cell organization to cell history, acknowledging that only cells that possessed mitochondria had the bioenergetic assets to achieve eukaryotic cell complexity, which explains why no in between beings existed in the prokaryote-to-eukaryote transition. This study attempts to approach the eukaryote origins from the standpoint of constructal theory, i.e., “For a flow system to persist in time (to survive), it must evolve in such a way that it provides easier and easier access to the currents that flow through it.” Therefore, the explanation is based on the direction of evolution in time, in which it has been introduced the thought that system structure should morph freely towards the optimal architecture and flow organization that minimize resistances to the internal flows (exergy losses) that are required for the system existence, which is proposed as the origin of eukaryotes, mitochondria, chloroplasts, and for the origin of the eukaryotic nucleus.

Elucidation of physiological functions of sphingolipid-related molecules by chemical approaches.

Sphingolipids (SLs), found in all animals, plants, and fungi and in certain prokaryotic organisms, exhibit essential physiological functions that cannot be replicated by other lipids. Although SLs and their related biomolecules behave as lipid mediators, skin barrier systems and epitopes, their detailed biological functions have not yet been revealed, unlike those of proteins and nucleic acids, because the biosynthesis of SLs is not governed by the central dogma. Recently, SLs have been widely studied in relation to diseases such as obesity, dementia, and neuron agenesis and have attracted attention as molecules related to unmet medical needs. This review presents the recent applications of the SL chemical biology in unmet medical needs.

Identification of Small RNAs in Streptomyces clavuligerus Using High-Resolution Transcriptomics and Expression Profiling During Clavulanic Acid Production.

Small non-coding RNAs play a pivotal role in regulating various metabolic processes in both prokaryotic and eukaryotic organisms. However, knowledge about small RNAs (sRNAs) in Streptomyces clavuligerus (S. clavuligerus) is scarce. This study aimed to use cutting-edge bioinformatics tools and a compendium of RNA-seq data to predict the potential coding of sRNAs that might be present in the genome of S. clavuligerus ATCC 27064. In the genome of S. clavuligerus, 606 intergenic regions (IGRs) are conserved, and 272 possess a highly thermodynamically stable and conserved secondary structure, indicating the presence of non-coding RNA in these regions. The transcriptome assembly of S. clavuligerus showed that the genome is completely functional, as all the annotated genes are expressed under the conditions analyzed. From this assembly, transcripts originating from IGRs were labeled as putative sRNAs, and their differential expression during the growth curve of S. clavuligerus for clavulanic acid (CA) production was established. The interactome of these differentially expressed (DE) RNAs displayed the sRNAs as global regulators, as they can have multiple mRNA targets. The functional annotation of the target genes of DE sRNAs demonstrated that they are directly involved in secondary metabolite production. Specifically, two sRNA have the genes of the biosynthetic gene cluster of CA as targets. Thus, these molecules add an additional layer to the regulatory cascade for CA biosynthesis, and we propose them as targets for metabolic engineering to increase CA production.

The Effect of a Photoactivated Ruthenium Nitrocomplex [RuCl(NO₂)(dppb)(4,4-2 Mebipy)] on the Viability of Eukaryotic and Prokaryotic Cells, Including Bacterial Biofilms

Antimicrobial resistance (AMR) is a critical global public health problem. Many bacterial pathogens use biofilm formation as their main pathogenicity mechanism, a practical tactic for surviving in natural settings and colonized host tissues. Research using ruthenium(II) complexes has demonstrated antibacterial action linked to photodynamic therapy, an alternate method of microbial control. Thus, in this work, the photosensitive nitro complex [RuCl(NO2)(dppb)(4,4-Mebipy)] (I) was prepared and the X-ray structure was determined. Then, we investigated the antibacterial and antibiofilm activities, antibiotic-associated effects, and cytotoxicity. The results showed that complex I exhibited promising antimicrobial activity with MIC values ranging from 4 to 256 µg/mL and MBC from 4 to 32 µg/mL. The antimicrobial activity of this nitro complex was significantly enhanced with blue light irradiation, as confirmed by agarose gel electrophoresis of the pBR322 DNA, which must be related to the DNA cleavage promoted by the photorelease of NO. A synergistic effect against Staphylococcus aureus and Staphylococcus epidermidis strains was observed when combined with ampicillin, which exhibited FICI values from 0.186 to 0.311. Interestingly, complex I associated with tetracycline showed a synergistic effect only on Escherichia coli. Regarding biofilms, the irradiated complex I showed antibacterial activity against biofilm formation and mature biofilms. Furthermore, SEM and confocal analyses revealed changes in cell morphology and damage to the wall and plasma membrane. Complex I presented a percentage of hemolysis between 2 and 4%, and no cytotoxic effect was observed against murine dermal fibroblasts. In conclusion, the photoactivated ruthenium(II) complex showed antibacterial and antibiofilm activity against relevant bacteria.

Optimized Construction of a Yeast SICLOPPS Library for Unbiased In Vivo Selection of Cyclic Peptides.

DNA-encoded libraries hold great potential for discovering small, cyclized peptides with drug potential. Split-intein circular ligation of peptides and proteins (SICLOPPS) is a well-established method for in vivo selection of cyclic peptides targeting specific intracellular components. However, the method has mainly been used in prokaryotic cells. In contrast, selection studies performed directly in eukaryotic cells allow for the identification of cyclic peptides promoting a functional outcome, without the need to define a specific cellular target. Here, we report the construction of a Saccharomyces cerevisiae-specific SICLOPPS library of 80 million members, via careful optimization of several steps to increase the size of the library. Individual library members were shown to be correctly expressed and processed in yeast. High-throughput sequencing was conducted on the randomized primer used for library construction and the pure yeast SICLOPPS library isolated from Escherichia coli. A distinct guanine insertion bias was observed in the peptide-encoding, randomized sequence, which was primarily attributed to the degenerate primer used to introduce the randomized sequence. Moreover, high-throughput sequencing was performed on the library before and after the induction of cyclic peptide expression in yeast. Importantly, expression of the SICLOPPS library in S. cerevisiae caused only a marginal further sequence bias. Our work paves the way for selection studies using a large and diverse library to identify cyclic peptides of therapeutic interest that promote a specific phenotypic outcome in eukaryotic organisms, with yeast representing a beneficial model system due to its high transformation efficiency.

Recent innovations in Baculovirus‐insect cell expression Systems for Therapeutic Protein Production

AbstractThe production of recombinant glycoproteins is pivotal for numerous therapies targeting human diseases, forming a cornerstone of the medical industry. The baculovirus‐insect cell system has emerged as a leading method for producing high‐value therapeutic glycoproteins. This system has several advantages, including rapid cultivation of insect cells, high production yields, and relatively straightforward post‐translational processing. Insect cells, being eukaryotic, have functional glycosylation pathways, offering a significant advantage over prokaryotic cells which lack these capabilities. These advantages are particularly promising for the expression and testing of post‐translational processing in de novo proteins, which are newly developed and extensively researched through AI‐based computational methods. However, challenges remain, particularly with expression levels and glycosylation modifications, which need to be addressed to optimize protein production. This review aims to provide a comprehensive overview of recent advancements in the baculovirus‐insect cell expression system, highlighting progress in insect cell utilization, expression optimization, glycosylation modifications, and the production of therapeutic proteins.

Reciprocal regulation between bacterial secretion systems and host metabolism: Enhancing bacterial intracellular survival capability.

Secretion systems are intricate nanomachines present on many bacterial cell membranes that deliver various bacterially-encoded effector proteins into eukaryotic or prokaryotic cells. They are pivotal in bacterial invasion, host colonization, and pathogenesis. After infection, bacteria employ these machines to deliver toxic effectors to the cytoplasm of host cells that disrupt their metabolic balance, such as interfering with glucose metabolism, promoting lipid droplets formation, altering amino acid profiles and mitochondrial morphology, and reducing ROS levels, to ensure bacterial intracellular survival. Furthermore, metabolites within host cells can modulate the expression and/or function of bacterial secretion systems. This review summarizes recent advancements in understanding the impact of bacterial secretion systems on host cell metabolism and the feedback regulation of host metabolites on these machines, providing novel perspectives on host-pathogen interactions and mechanisms of bacterial pathogenesis.

Interplay of niche and respiratory network in shaping bacterial colonization.

The human body is an intricate ensemble of prokaryotic and eukaryotic cells, and this coexistence relies on the interplay of many biotic and abiotic factors. The inhabiting microbial population has to maintain its physiological homeostasis under highly dynamic and often hostile host environments. While bacterial colonization primarily relies on the metabolic suitability for the niche, there are reports of active remodeling of niche microenvironments to create favorable habitats, especially in the context of pathogenic settlement. Such physiological plasticity requires a robust metabolic system, often dependent on an adaptable energy metabolism. This review focuses on the respiratory electron transport system and its adaptive consequences within the host environment. We provide an overview of respiratory chain plasticity, which allows pathogenic bacteria to niche-specify, niche-diversify, mitigate inflammatory stress, and outcompete the resident microbiota. We have reviewed existing and emerging knowledge about the role of respiratory chain components responsible for the entry and exit of electrons in influencing the pathogenic outcomes.

Cave Pools in Carlsbad Caverns National Park Contain Diverse Bacteriophage Communities and Novel Viral Sequences

Viruses are the most abundant biological entities on Earth, and they play a critical role in the environment and biosphere where they regulate microbial populations and contribute to nutrient cycling. Environmental viruses have been the most studied in the ocean, but viral investigations have now spread to other environments. Here, viral communities were characterized in four cave pools in Carlsbad Caverns National Park to test the hypotheses that (i) viral abundance is ten-fold higher than prokaryotic cell abundance in cavern pools, (ii) cavern pools contain novel viral sequences, and (iii) viral communities in pools from developed portions of the cave are distinct from those of pools in undeveloped parts of the same cave. The relationship between viral and microbial abundance was determined through direct epifluorescence microscopy counts. Viral metagenomes were constructed to examine viral diversity among pools, identify novel viruses, and characterize auxiliary metabolic genes (AMGs). Bacterial communities were characterized by 16S rRNA gene amplicon sequencing. Epifluorescence microscopy showed that the ratio of viral-like particles (VLPs) to microorganisms was approximately 22:1 across all sites. Viral communities from pools with higher tourist traffic were more similar to each other than to those from less visited pools, although surprisingly, viruses did not follow the same pattern as bacterial communities, which reflected pool geochemistry. Bacterial hosts predicted from viral sequences using iPHoP showed overlap with both rare and abundant genera and families in the 16S rRNA gene dataset. Gene-sharing network analysis revealed high viral diversity compared to a reference viral database as well as to other aquatic environments. AMG presence showed variation in metabolic potential among the four pools. Overall, Carlsbad Cavern harbors novel viruses with substantial diversity among pools within the same system, indicating that caves are likely an important repository for unexplored viromes.

Maternal microbiome and the impact in pre- and post-natal life

The maternal organism, including the microbiome of the oral cavity, vaginal and intestinal microbiome, influences the risk of pregnancy outcomes related to fetal health even after birth. Recent research reveals that it is not only the vaginal cavity, blood circulation, placenta or amniotic fluid that contain bacteria. While the interaction of bacteria or their metabolites with the fetus or newborn has serious consequences by causing diseases or malformations. An alternative for women undergoing cesarean section is vaginal seeding, a practice that involves transferring vaginal fluids into the body of the newborn. The purpose is to provide beneficial maternal bacteria to babies. Moreover, factors such as probiotics, oral hygiene and nutrition have been shown to influence the microbiome. The consumption of antibiotics during the gestational stage could lead to preterm delivery due to deficiency of the maternal gut microbiome. Some hypotheses suggest alterations in neurodevelopment, for example, in thalamocortical axonogenesis, where its deficiency is related to decreased limb sensitivity. Research on the bacterial 16S rRNA sequence comparison makes it possible to establish phylogenetic relationships between prokaryotic organisms; this new identification method uses the 16S rRNA polyribonucleotide, makes it possible to distinguish individual bacteria, especially in the case of difficult ones. It consists of 3 steps: gene amplification from the sample, nucleotide sequence determination, and sequence analysis. As a result, it has been discovered that the fetus does not grow in a sterile environment.

Design of a Bioluminescent Assay Platform for Quantitative Measurement of Histone Acetyltransferase Enzymatic Activity.

Protein acetylation and acylation are widespread post-translational modifications (PTMs) in eukaryotic and prokaryotic organisms. Histone acetyltransferase (HATs) enzymes catalyze the addition of short-chain acyl moieties to lysine residues on cellular proteins. Many HAT members are found to be dysregulated in human diseases, especially oncological processes. Screening potent and selective HAT inhibitors has promising application for therapeutic innovation. A biochemical assay for quantification of HAT activity utilizing luminescent output is highly desirable to improve upon limitations associated with the classic radiometric assay formats. Here we report the design of a bioluminescent technological platform for robust and sensitive quantification of HAT activity. This platform utilizes the metabolic enzyme acetyl-CoA synthetase 1 (ACS1) for a coupled reaction with firefly luciferase to generate luminescent signal relative to the HAT-catalyzed acetylation reaction. The biochemical assay was implemented in microtiter plate format and our results showed this assay sensitively detected catalytic activity of HAT enzyme p300, accurately measured its steady-state kinetic parameters of histone acetylation and measured the inhibitory potency of HAT inhibitor. This platform demonstrated excellent robustness, reproducibility, and signal-to-background ratios, with a screening window Z'=0.79. Our new bioluminescent design provides an alternative means for HAT enzymatic activity quantitation and HAT inhibitor screening.

Identity confirmation of extracellular vesicles by capillary electrophoresis using non-specific dyes

Extracellular vesicles (EVs) are membrane-enveloped nanostructures secreted by prokaryotic and eukaryotic cells. Carrying a variety of essential components, EVs are involved in numerous biological processes including intercellular communication and immunological response. The characterization of extracellular vesicles with capillary electrophoresis (CE) faces several challenges. Due to the limited availability of commercial standards, the identity confirmation of the EV signal is among these issues. In the presented work, a method for identity confirmation of EVs by CE was developed. Identification was based on the application of non-specific dyes and fluorescence detection, and EVs obtained from Escherichia coli and citrus fruits (Citrus limon and Citrus aurantifolia) were used. It was found that the presented methodology effectively detects the cargo carried by EVs (including nucleic acids, proteins, and lipids) and enables the staining of subpopulations separated by CE. The strategy holds promise to be utilized for EV heterogeneity assessment.

In vitro Production and Characterization of a Recombinant Viral Protein (rVP1) of Coxsackievirus B3 (CVB3): Contribution for the Development of a Subunit Vaccine or an Immunodiagnostic Reagent

Coxsackievirus B3 is an Enterovirus implicated in diverse human pathologies, from viral myocarditis to neurological disorders. There isn’t a medicinal agent or vaccine for CVB3 in clinical use at the moment, despite the possibility that vaccination could lower the prevalence of these illnesses. This study focuses on the in vitro production and characterization of the viral protein 1 (VP1) in the objective to use it as subunit vaccine and/or immunodiagnostic reagent. VP1 is considered as the most immunogenic capsid protein of the CVB3 surface. We amplified the VP1 whole gene by RT-PCR from the extracted wild type Nancy strain RNA, then cloned it into the pUC19 plasmid expression vector, and expressed it in E. coli DH5a prokaryotic cells. The obtained recombinant proteins were then analyzed by SDS-PAGE and characterized by Bioinformatic software tools. Our results revealed that the produced recombinant amino acid VP1 (rVP1) is highly identical to the VP1 of the CVB3 wild-type strain and has very similar physicochemical properties. In addition, we demonstrated that rVP1 has the highest number of phosphorylation sites which means that rVP1 can translate the host cell signal via the phosphorylation mechanism. Moreover, The Linear B cell epitope analysis showed that the rVP1 contains many epitope regions that should be recognized by the humoral host immune response. Taken together, results demonstrate that the cloned and recombined expressed viral protein could be used to carry out any studies concerning the development a protein subunit vaccine against CVB3 infections or an immunodiagnostic reagent for detecting the virus in samples.

The High Millimolar Concentration of ATP: A Fundamental & Foundational Feature of Eukaryotic, Archaeotic, and Prokaryotic Domains.

Measurement of the adenosine triphosphate (ATP) concentration among different cells, tissues and organs and even across the phylogenetic tree ordinarily yields exceedingly high concentrations at the millimolar (mM) level. This represents a conundrum in that ATP-driven cellular functions only require micromolar (μM) values. Considering that nature is ordinarily conservative in the generation of high-energy phosphatic metabolites such as ATP, a potential major role for ATP has been completely overlooked and may be of paramount importance because ATP is a hydrotrope. In all phylogenetic domains, reports have established that the excessively high mM concentration of ATP is present in studies of eukaryotic cellular and tissue homogenates, living tissues, and a living organ as well as archaeotic and prokaryotic organisms. These ATP concentrations are also present in contemporary relatives of microorganisms having progenitors existing in the Precambrian Era. This feature is fundamental to cell biology across taxonomic domains. These features are interpreted as serving a foundational molecular function for maintaining organismal homeostasis. We hypothesize that ATP prevents pathological protein aggregation and maintains protein solubility through its hydrotropic feature in cells, tissues, and organs.