Breakdown Pathway Research Articles

Activation of persulfate by MOF-derived MnFeOx to efficiently degrade sulfadiazine: Synergistic effects from free radicals and singlet oxygen

In this study, a facile hydrothermal approach was used to synthesize the MOFs materials [Mn-doped MIL-101(Fe)]. These materials were subsequently subjected to high temperature annealing process via calcination to generate the MOFs derivatives (MnFeOx). The synthesized materials were utilized for the removal of sulfadiazine (SDZ) removal through activated peroxymonosulfate (PMS). At the optimum Mn doping, the removal ratio of SDZ could reach 98.9 % (at natural pH) within 20 min. In addition, the study investigated the influence of PMS dosage, catalyst dosage, initial pH, various inorganic anions, and humic acid (HA) on the degradation of SDZ. The activation processes of both free and non-free radicals in the MnFeOx/PMS system were examined by free radical quenching experiments and electron paramagnetic resonance (EPR) techniques. The analysis of the degradation products of SDZ was conducted by using high performance liquid chromatography-mass spectrometry (HPLC-MS), and the breakdown pathways of SDZ in the MnFeOx/PMS system were proposed.

Chitosan in biomedicine: A comprehensive review of recent developments

A naturally occurring biopolymer, chitosan has found widespread application in the fields of biomedical research and healthcare enhancement. Through a thorough analysis of recent research findings and technological advancements, this review seeks to provide a comprehensive overview of the latest developments and potential uses of chitosan in the field of biomedicine. Microbe cells' anionic lipopolysaccharides and surface proteins interact with the polycationic composition, releasing intracellular components and resulting in cell death. The degree of antimicrobial activity is highly dependent on the particular type of microbe being targeted. Chitosan is highly susceptible to enzymatic hydrolytic breakdown pathways in the human body, especially when lysozymes are involved. Moreover, adding chitosan derivatives to mice's diet improved their health by increasing their enzymatic activity, which is related to its scavenging properties and shows anti-microbial activity. Furthermore, the assessment of sutures manufactured from this substance emphasizes their noteworthy input to the field of modern wound care.

Enhancing cellulose acetate biodegradability in cigarette filters: an in-depth analysis of thermal alkaline pretreatment, microbial dynamics, and breakdown pathway prediction

BackgroundThe demand for bioplastics has increased exponentially as they have emerged as alternatives to petrochemical plastics. However, there is a substantial lack of knowledge regarding bioplastic degradation. This study developed a novel pretreatment method to improve the accessibility of a bioplastic substrate for biodegradation. In this study, cellulose acetate, a bioplastic found in the world’s most littered waste, e.g. cigarette filters, was selected as a potential substrate. Before anaerobic digestion, three thermal alkaline pretreatments: TA 30 °C, TA 90 °C, and TA 121 °C, were used to evaluate their effects on the chemical alterations of cellulose acetate.ResultThe ester groups in cellulose acetate were significantly reduced by the TA 30 °C pretreatment, as seen by a decrease in C = O stretching vibrations and shortening of C − O stretches (1,270 ∼ 1,210 cm− 1), indicating effective removal of acetyl groups. This pretreatment significantly enhanced cellulose acetate biodegradability to a maximum of 91%, surpassing the previously reported cellulose acetate degradation. Methane production increased to 695.0 ± 4 mL/g of volatile solid after TA 30 °C pretreatment, indicating enhanced cellulose acetate accessibility to microorganisms, which resulted in superior biogas production compared to the control (306.0 ± 10 mL/g of volatile solid). Diverse microbes in the anaerobic digestion system included hydrolytic (AB240379_g, Acetomicrobium, FN436103_g, etc.), fermentative, and volatile fatty acids degrading bacteria (JF417922_g, AB274492_g, Coprothermobacter, etc.), with Methanobacterium and Methanothermobacter being the sole hydrogenotrophic methanogens in the anaerobic digestion system. Additionally, an attempt to predict the pathway for the effective degradation of cellulose acetate from the microbial community in different pretreatment conditions.ConclusionsTo the best of our knowledge, this is the first study to estimate the maximum cellulose acetate degradation rate, with a simple and cost-effective pretreatment procedure. This approach holds promise for mitigating the environmental impact of cellulose acetate of cigarette filters and presents a sustainable and economically viable waste management strategy.

Detection of Sulfoquinovosidase Activity in Cell Lysates Using Activity‐Based Probes

AbstractThe sulfolipid sulfoquinovosyl diacylglycerol (SQDG), produced by plants, algae, and cyanobacteria, constitutes a major sulfur reserve in the biosphere. Microbial breakdown of SQDG is critical for the biological utilization of its sulfur. This commences through release of the parent sugar, sulfoquinovose (SQ), catalyzed by sulfoquinovosidases (SQases). These vanguard enzymes are encoded in gene clusters that code for diverse SQ catabolic pathways. To identify, visualize and isolate glycoside hydrolase CAZY‐family 31 (GH31) SQases in complex biological environments, we introduce SQ cyclophellitol‐aziridine activity‐based probes (ABPs). These ABPs label the active site nucleophile of this enzyme family, consistent with specific recognition of the SQ cyclophellitol‐aziridine in the active site, as evidenced in the 3D structure of Bacillus megaterium SQase. A fluorescent Cy5‐probe enables visualization of SQases in crude cell lysates from bacteria harbouring different SQ breakdown pathways, whilst a biotin‐probe enables SQase capture and identification by proteomics. The Cy5‐probe facilitates monitoring of active SQase levels during different stages of bacterial growth which show great contrast to more traditional mRNA analysis obtained by RT‐qPCR. Given the importance of SQases in global sulfur cycling and in human microbiota, these SQase ABPs provide a new tool with which to study SQase occurrence, activity and stability.

Detection of Sulfoquinovosidase Activity in Cell Lysates Using Activity-Based Probes.

The sulfolipid sulfoquinovosyl diacylglycerol (SQDG), produced by plants, algae, and cyanobacteria, constitutes a major sulfur reserve in the biosphere. Microbial breakdown of SQDG is critical for the biological utilization of its sulfur. This commences through release of the parent sugar, sulfoquinovose (SQ), catalyzed by sulfoquinovosidases (SQases). These vanguard enzymes are encoded in gene clusters that code for diverse SQ catabolic pathways. To identify, visualize and isolate glycoside hydrolase CAZY-family 31 (GH31) SQases in complex biological environments, we introduce SQ cyclophellitol-aziridine activity-based probes (ABPs). These ABPs label the active site nucleophile of this enzyme family, consistent with specific recognition of the SQ cyclophellitol-aziridine in the active site, as evidenced in the 3D structure of Bacillus megaterium SQase. A fluorescent Cy5-probe enables visualization of SQases in crude cell lysates from bacteria harbouring different SQ breakdown pathways, whilst a biotin-probe enables SQase capture and identification by proteomics. The Cy5-probe facilitates monitoring of active SQase levels during different stages of bacterial growth which show great contrast to more traditional mRNA analysis obtained by RT-qPCR. Given the importance of SQases in global sulfur cycling and in human microbiota, these SQase ABPs provide a new tool with which to study SQase occurrence, activity and stability.

Degradation and inhibition kinetics of phenanthrene by Alcaligenes ammonioxydans, [VITRPS2] strain isolated from petroleum-contaminated soil

Phenanthrene, a common three-ring polyaromatic hydrocarbon [PAH], originates from sources like grilled meals, tobacco, crude oil, coal tar, and automobile exhaust. Recognized as a hazardous PAH, it is often targeted for bioremediation due to its sustainability and potential for full mineralization. In this study, we focus on biodegrading phenanthrene using the strain Alcaligenes ammonioxydans [VITRPS2], isolated from petroleum-contaminated soil. At 5 mg/ml, degradation occurred at a rate constant of 0.0181/day, with half-life values of 2.7 and 4.49 according to first and second-order kinetics, respectively. Employing a one-factor-at-a-time [OFAT] approach, we optimized biodegradation conditions within Luria–Bertani [LB] media. Under optimal conditions—pH 8.0, 8% inoculum concentration, and 37 °C incubation over seven days—the strain achieved maximal growth with phenanthrene as the sole carbon source. It exhibited a degradation efficiency of up to 72% for phenanthrene under these conditions. Gas chromatography-mass spectrometry [GC–MS] analysis revealed principal metabolites of the breakdown pathway, including salicylic acid, catechol, and various phthalic acid derivatives. This underscores the strain's potential for remediating environments polluted by PAH metabolites, showcasing its remarkable capability for complete phenanthrene degradation.Graphical abstract

Glycerol metabolism impacts biofilm phenotypes and virulence in Pseudomonas aeruginosa via the Entner-Doudoroff pathway.

Pseudomonas aeruginosa is a ubiquitous bacterium and a notorious opportunistic pathogen that forms biofilm structures in response to many environmental cues. Biofilm formation includes attachment to surfaces and the production of the exopolysaccharide Pel, which is present in both the PAO1 and PA14 laboratory strains of P. aeruginosa. Biofilms help protect bacterial cells from host defenses and antibiotics and abet infection. The carbon source used by the cells also influences biofilm, but these effects have not been deeply studied. We show here that glycerol, which can be liberated from host surfactants during infection, encourages surface attachment and magnifies colony morphology differences. We find that glycerol kinase is important but not essential for glycerol utilization and relatively unimportant for biofilm behaviors. Among downstream enzymes predicted to take part in glycerol utilization, Edd stood out as being important for glycerol utilization and for enhanced biofilm phenotypes in the presence of glycerol. Thus, gluconeogenesis and catabolism of anabolically produced glucose appear to impact not only the utilization of glycerol but also glycerol-stimulated biofilm phenotypes. Finally, waxworm moth larvae and nematode infection models reveal that interruption of the Entner-Doudoroff pathway, but not abrogation of glycerol phosphorylation, unexpectedly increases P. aeruginosa lethality in both acute and chronic infections, even while stimulating a stronger immune response by Caenorhabditis elegans.IMPORTANCEPseudomonas aeruginosa, the ubiquitous environmental bacterium and human pathogen, forms multicellular communities known as biofilms in response to various stimuli. We find that glycerol, a common carbon source that bacteria can use for energy and biosynthesis, encourages biofilm behaviors such as surface attachment and colony wrinkling by P. aeruginosa. Glycerol can be derived from surfactants that are present in the human lungs, a common infection site. Glycerol-stimulated biofilm phenotypes do not depend on phosphorylation of glycerol but are surprisingly impacted by a glucose breakdown pathway, suggesting that it is glycerol utilization, and not its mere presence or cellular import, that stimulates biofilm phenotypes. Moreover, the same mutations that block glycerol-stimulated biofilm phenotypes also impact P. aeruginosa virulence in both acute and chronic animal models. Notably, a glucose-breakdown mutant (Δedd) counteracts biofilm phenotypes but shows enhanced virulence and stimulates a stronger immune response in Caenorhabditis elegans.

Plant glucosinolate biosynthesis and breakdown pathways shape the rhizosphere bacterial/archaeal community.

Rhizosphere microbial community assembly results from microbe-microbe-plant interactions mediated by small molecules of plant and microbial origin. Studies with Arabidopsis thaliana have indicated a critical role of glucosinolates in shaping the root and/or rhizosphere microbial community, likely through breakdown products produced by plant or microbial myrosinases inside or outside of the root. Plant nitrile-specifier proteins (NSPs) promote the formation of nitriles at the expense of isothiocyanates upon glucosinolate hydrolysis with unknown consequences for microbial colonisation of roots and rhizosphere. Here, we generated the A. thaliana triple mutant nsp134 devoid of nitrile formation in root homogenates. Using this line and mutants lacking aliphatic or indole glucosinolate biosynthesis pathways or both, we found bacterial/archaeal alpha-diversity of the rhizosphere to be affected only by the ability to produce aliphatic glucosinolates. In contrast, bacterial/archaeal community composition depended on functional root NSPs as well as on pathways of aliphatic and indole glucosinolate biosynthesis. Effects of NSP deficiency were strikingly distinct from those of impaired glucosinolate biosynthesis. Our results demonstrate that rhizosphere microbial community assembly depends on functional pathways of both glucosinolate biosynthesis and breakdown in support of the hypothesis that glucosinolate hydrolysis by myrosinases and NSPs happens before secretion of products to the rhizosphere.

Anaerobic heme recycling by gut microbes: Important methods for monitoring porphyrin production.

Heme is the most abundant species of iron inside the human body and an essential cofactor for numerous electron/chemical group transfer reactions and catalyses, especially those involving O2. Whole anaerobic biomes exist that also depend on heme but lack widespread, O2-dependent pathways for heme synthesis and breakdown. The gastrointestinal tract is an anaerobic ecosystem where many microbes are auxotrophic for heme, and where the abundant members of the Bacteroidetes phylum convert heme into iron and porphyrins. Working with mixtures of these hydrophobic compounds presents challenges for analyses, especially when their source is biological. In this brief chapter, we detail a handful of important methods and point out caveats necessary for their concurrent detection, separation, and quantification.

Chapter 11 - White matter abnormalities in amino acid disorders and organic acidurias

Inborn errors of metabolism (IEMs) are traditionally the domain of pediatricians and internists for metabolic diseases. In general, neurologists only become involved when these disorders are complicated by neurologic symptoms such as seizures, developmental delay, or motor problems. However, in recent years and mainly due to the successes of next-generation sequencing, the number of IEMs primarily presenting with neurologic symptoms and not detected by classic biochemical testing has grown significantly. This in particular relates to disorders in the biosynthesis of amino acids. Therefore, I will start by discussing defects in the synthesis pathways of the amino acids serine, glutamine, proline, and asparagine. In these disorders, the amino acid can be low in body fluids with biochemical testing, but more frequently are completely normal and although are in different metabolic pathways, they share many clinical features such as hypomyelination and white matter abnormalities. Next, I will discuss classic amino acid disorders and organic acid disorders due to defects in breakdown pathways characterized by elevations of key metabolites in body fluids and associated with neurologic abnormalities and white matter changes on MRI.

Characterization of a phenotypically severe animal model for human AB-Variant GM2 gangliosidosis

AB-Variant GM2 gangliosidosis (ABGM2) is a rare and lethal genetic disorder caused by mutations in the GM2A gene that lead to fatal accumulation of GM2 gangliosides (GM2) in neurons of the central nervous system (CNS). GM2A encodes a transport protein known as GM2 activator (GM2A) protein, which is essential for degrading GM2 into their GM3 form. ABGM2 presents in infantile-, juvenile-, and adult-onset forms; of the three, the infantile-onset is the most prominent, and by far the most severe, as evidenced by high levels of GM2 accumulation, widespread neurodegeneration, and death by the age of 4. Gm2a−/− mice are commonly used as a model of ABGM2. These mice are characterized by phenotypes most representative of predicted adult-onset form of ABGM2, which include moderate GM2 accumulation and mild neurological defects. This mild phenotype has been attributed to compensation by alternative GM2 degradation pathways mediated by sialidase, neuraminidase 3 (NEU3), a pathway that is more prominent in mice than humans. To assess the extent to which NEU3 contributes to GM2 degradation, we generated double knock-out (Gm2a−/−Neu3−/−) mice. Compellingly, these mice present with a clinical phenotype resembling that of a more severe ABGM2, including ataxia, reduced mobility and coordination, weight loss, poor body scores, and lethality by 6–7 months. Furthermore, these phenotypes correlate with a dramatic increase in GM2 accumulation in the CNS compared to levels observed in either Gm2a−/− or Neu3−/− mice. Taken together, these studies, for the first-time, confirm that the mild neurological phenotype of Gm2a−/− mice is due to compensatory activity on GM2 catabolism through an alternate breakdown pathway involving NEU3. These studies support the use of double knockout mice as a novel and highly relevant model for pre-clinical drug studies in a more severe form of ABGM2.

Elucidating microbial mechanisms of microcystin-LR degradation in Lake Erie beach sand through metabolomics and metatranscriptomics

As one of five Laurentian Great Lakes, Lake Erie ranks among the top freshwater drinking sources and ecosystems globally. Historical and current agriculture mismanagement and climate change sustains the environmental landscape for late summer cyanobacterial harmful algal blooms, and consequently, cyanotoxins such as microcystin (MC). Microcystin microbial degradation is a promising mitigation strategy, however the mechanisms controlling the breakdown of MCs in Lake Erie are not well understood. Pelee Island, Ontario, Canada is located in the western basin of Lake Erie and the bacterial community in the sand has demonstrated the capacity of metabolizing the toxin. Through a multi-omic approach, the metabolic, functional and taxonomical signatures of the Pelee Island microbial community during MC-LR degradation was investigated over a 48-hour period to comprehensively study the degradation mechanism. Cleavage of bonds surrounding nitrogen atoms and the upregulation of nitrogen deamination (dadA, alanine dehydrogenase, leucine dehydrogenase) and assimilation genes (glnA, gltB) suggests a targeted isolation of nitrogen by the microbial community for energy production. Methylotrophic pathways RuMP and H4MPT control assimilation and dissimilation of carbon, respectively and differential abundance of Methylophilales indicates an interconnected role through electron exchange of denitrification and methylotrophic pathways. The detected metabolites did not resolve a clear breakdown pathway, but rather the diversity of products in combination with taxonomic and functional results supports that a variety of strategies are applied, such as epoxidation, hydroxylation, and aromatic degradation. Annual repeated exposure to the toxin may have allowed the community to adaptatively establish a novel pathway through functional plasticity and horizontal gene transfer. The culmination of these results reveals the complexity of the Pelee Island sand community and supports a dynamic and cooperative metabolism between microbial species to achieve MC degradation.

Microbial degradation of plant toxins.

Plants produce a variety of secondary metabolites in response to biotic and abiotic stresses. Although they have many functions, a subclass of toxic secondary metabolites mainly serve plants as deterring agents against herbivores, insects, or pathogens. Microorganisms present in divergent ecological niches, such as soil, water, or insect and rumen gut systems have been found capable of detoxifying these metabolites. As a result of detoxification, microbes gain growth nutrients and benefit their herbivory host via detoxifying symbiosis. Here, we review current knowledge on microbial degradation of toxic alkaloids, glucosinolates, terpenes, and polyphenols with an emphasis on the genes and enzymes involved in breakdown pathways. We highlight that the insect-associated microbes might find application in biotechnology and become targets for an alternative microbial pest control strategy.

Chemical stability investigation of haloolefin refrigerants and their blends with lubricants

This paper examines the findings of highly accelerated life tests (HALT), conducted according to ASHRAE Standard 97 sealed glass tube methodology for R-123 like, R-134a like and R-410A/R-404A like lower global warming potential (GWP) alternatives. Additionally, possible chemical breakdown pathways are reviewed for the new haloolefin refrigerants. Note, compositions for the following blends are shown as percent by weight. For R-123 like refrigerants, R-1336mzz(Z), R-1336mzz(E), R-514A (74.7% R-1336mzz(Z)/25.3% R-1130(E)), R-1233zd(E), and R-1224yd(Z) were examined with and without lubricants. R-1233zd(E) and R-1224yd(Z) were evaluated with mineral oil and the rest were evaluated with polyalkylene glycol (PAG), polyolester (POE) and polyvinyl ether (PVE) lubricants. For R-134a like refrigerants, R-1234ze(E), R-450A (42% R-134a/58% R-1234ze(E)), R-515B (91.1% R-1234ze(E)/8.9% R-227ea), R-1234yf, R-513A (56% R-1234yf/44% R-134a), and R-516A (77.5% R-1234yf/8.5% R-134a/14.0% R-152a) were examined with and without PAG, POE, and PVE lubricants. For R-410A/R-404A like, R-454B (68.9% R-32/31.1% R-1234yf), R-454C (21.5% R-32/78.5% R-1234yf), R-455A (3.0% R-744/21.5% R-32/75.5% R-1234yf), and R-468A (3.5% R-1132a/21.5% R-32/75.0% R-1234yf) were also examined with PAG, POE and PVE lubricants. R-466A (49% R-32/11.5% R-125/39.5% CF3I) was part of this study but will not be covered since it does not contain a haloolefin component.

Rice Bran Supplementation Ameliorates Gut Dysbiosis and Muscle Atrophy in Ovariectomized Mice Fed with a High-Fat Diet.

Rice bran, a byproduct of rice milling, is rich in fiber and phytochemicals and confers several health benefits. However, its effects on gut microbiota and obesity-related muscle atrophy in postmenopausal status remain unclear. In this study, we investigated the effects of rice bran on gut microbiota, muscle synthesis, and breakdown pathways in estrogen-deficient ovariectomized (OVX) mice receiving a high-fat diet (HFD). ICR female mice were divided into five groups: sham, OVX mice receiving control diet (OC); OVX mice receiving HFD (OH); OVX mice receiving control diet and rice bran (OR); and OVX mice receiving HFD and rice bran (OHR). After twelve weeks, relative muscle mass and grip strength were high in rice bran diet groups. IL-6, TNF-α, MuRf-1, and atrogin-1 expression levels were lower, and Myog and GLUT4 were higher in the OHR group. Rice bran upregulated the expression of occludin and ZO-1 (gut tight junction proteins). The abundance of Akkermansiaceae in the cecum was relatively high in the OHR group. Our finding revealed that rice bran supplementation ameliorated gut barrier dysfunction and gut dysbiosis and also maintained muscle mass by downregulating the expression of MuRf-1 and atrogin-1 (muscle atrophy-related factors) in HFD-fed OVX mice.

Pentachlorophenol-induced hemotoxicity diminishes antioxidant potential and oxidizes proteins, thiols, and lipids in rat blood: An in vivo study

Pentachlorophenol (PCP) is an excessively used wood preservative and pesticide, which has resulted in human exposure raising concerns about its potential toxic effects. This study is designed to evaluate the hemotoxicity of PCP in adult rats. Wistar rats were orally administered PCP (25–150 mg/kg bw) for five days while untreated (control) rats received corn oil. Animals were sacrificed, blood was taken and fractionated into plasma and red blood cells (RBC). PCP administration increased methemoglobin formation but decreased methemoglobin reductase activity. Significantly increased hydrogen peroxide level indicates initiation of oxidative stress condition in blood. PCP increased the oxidation of thiols, proteins and lipids, lowered glutathione levels, and compromised the antioxidant status of RBC in treated rats. Enzymes of the pathways of glucose breakdown, glycolysis and phosphogluconate pathway, were inhibited. Markers of liver damage were increased in the plasma of PCP-treated rats suggesting hepatotoxicity. This was confirmed by histopathological analysis of stained liver sections. Activity of xanthine oxidase, a reactive oxygen species (ROS) generating pro-oxidant enzyme, was increased. These hematological changes could be a result of the increased generation of ROS or direct chemical transformation by transient reaction species. These results show that PCP induces redox imbalance, diminishes antioxidant potential, inhibits metabolic pathways, and oxidizes cellular components in rat blood.This study suggests an elaborated possible molecular mechanism of PCP toxicity, and similar compounds so that methods can be devised to minimize its damaging effect.

Strikingly conserved gene expression changes of polyamine regulating enzymes among various forms of acute and chronic kidney injury

The polyamines spermidine and spermine and their common precursor molecule putrescine are involved in tissue injury and repair. Here, we test the hypothesis that impaired polyamine homeostasis contributes to various kidney pathologies in mice during experimental models of ischemia-reperfusion, transplantation, rhabdomyolysis, cyclosporine treatment, arterial hypertension, diabetes, unilateral ureteral obstruction, high oxalate feeding, and adenine-induced injuries. We found a remarkably similar pattern in most kidney pathologies with reduced expression of enzymes involved in polyamine synthesis together with increased expression of polyamine degrading enzymes. Transcript levels of amine oxidase copper-containing 1 (Aoc1), an enzyme which catalyzes the breakdown of putrescine, were barely detectable by in situ mRNA hybridization in healthy kidneys. Aoc1 was highly expressed upon various experimental kidney injuries resulting in a significant reduction of kidney putrescine content. Kidney levels of spermine were also significantly reduced, whereas spermidine was increased in response to ischemia-reperfusion injury. Increased Aoc1 expression in injured kidneys was mainly accounted for by an Aoc1 isoform that harbors 22 additional amino acids at its N-terminus and shows increased secretion. Mice with germline deletion of Aoc1 and injured kidneys showed no decrease of kidney putrescine content; although they displayed no overt phenotype, they had fewer tubular casts upon ischemia-reperfusion injury. Hyperosmotic stress stimulated AOC1 expression at the transcriptional and post-transcription levels in metanephric explants and kidney cell lines. AOC1 expression was also significantly enhanced after kidney transplantation in humans. These data demonstrate that the kidneys respond to various forms of injury with down-regulation of polyamine synthesis and activation of the polyamine breakdown pathway. Thus, an imbalance in kidney polyamines may contribute to various etiologies of kidney injury.

A D-2-hydroxyglutarate dehydrogenase mutant reveals a critical role for ketone body metabolism in Caenorhabditis elegans development.

In humans, mutations in D-2-hydroxyglutarate (D-2HG) dehydrogenase (D2HGDH) result in D-2HG accumulation, delayed development, seizures, and ataxia. While the mechanisms of 2HG-associated diseases have been studied extensively, the endogenous metabolism of D-2HG remains unclear in any organism. Here, we find that, in Caenorhabditis elegans, D-2HG is produced in the propionate shunt, which is transcriptionally activated when flux through the canonical, vitamin B12-dependent propionate breakdown pathway is perturbed. Loss of the D2HGDH ortholog, dhgd-1, results in embryonic lethality, mitochondrial defects, and the up-regulation of ketone body metabolism genes. Viability can be rescued by RNAi of hphd-1, which encodes the enzyme that produces D-2HG or by supplementing either vitamin B12 or the ketone bodies 3-hydroxybutyrate (3HB) and acetoacetate (AA). Altogether, our findings support a model in which C. elegans relies on ketone bodies for energy when vitamin B12 levels are low and in which a loss of dhgd-1 causes lethality by limiting ketone body production.

457 CD39 Expression in the Glioblastoma Microenvironment does not Affect T Cell Exhaustion or Survival

INTRODUCTION: Glioblastoma (GBM), the most common malignant brain tumor, is notable for its dysfunctional T-cells. The initial optimism of Immune checkpoint inhibitors has been tempered as these agents have not been successful in treating GBM. CD39 is the first enzyme in the breakdown of ATP to adenosine and is highly upregulated in GBM. CD39 is also thought to contribute to T-cell exhaustion. However, inhibiting the CD39 ATP breakdown pathway did not improve survival in mice, even with PD-1 inhibition, leading to uncertainty about whether non-enzymatic functions of CD39 give it a role in T-cell exhaustion in GBM. METHODS: We performed intracranial injections of SB28 and GL261 GBM cell lines into CD39 global nulls (KO) and C57BL/6J (WT) mice and measured survival. Another cohort of mice were similarly injected and sacrificed after 2 weeks. These tumors were analyzed for T-cell exhaustion markers using flow cytometry. RESULTS: There was no survival benefit seen in either cell line when compared to wild type mice. The immune cell profile was not found to be statistically different between KO and WT mice. Next, expression of known exhaustion markers such as PD-1, TIM3, and LAG3 on tumor infiltrating lymphocytes (TILs) were analyzed. Expression of TIM3 and 2B4 were found to be increased in CD39KO TILs compared to WT (p < 0.01). Additionally, numerous exhaustion markers such as LAG3, TIGIT, and CTLA4 were decreased on TILs compared to peripheral blood T-cells in tumor-bearing mice (p < 0.01). CONCLUSIONS: These results suggest that CD39 lacks a direct impact on GBM outcomes but may affect the expression of other exhaustion markers. Further studies will continue to investigate the interaction between immune checkpoint molecules in GBM.

The photolytic conversion of 4-nonylphenol to 4-nonylcatechol within snowpack of the Palisade Glacier, Sierra Nevada, CA, USA

4-Nonylphenol (4-NP), an environmental pollutant with potent ecotoxicological effects, has been discovered in significant quantities in glacial ice and snow of the Sierra Nevada Mountain Range, CA. Photolysis of 4-NP is suspected to be a major, if not the sole, breakdown pathway in snow. However, the photolysis process has yet to be characterized in detail for this unique environment. This study therefore seeks to (1) confirm the presence of the major photolysis product within snowpack and snowmelt samples from the Palisade Glacier, CA, (2) determine key photolysis parameters through laboratory assays in snow analogs, and (3) compute environmentally relevant photolysis rates in snowpack via a spectral solar irradiance model parameterized for the Palisade Glacier. The primary photooxidation product of 4-NP, 4-nonylcatechol (4-NC), was synthesized and characterized by NMR and GC–MS for use as a reference standard in the detection of 4-NC in environmental samples. 4-NP was detected in all snowpack (n = 4) and snowmelt (n = 5) samples, with concentrations of 1.05 (± 0.11) μg L−1 and 1.28 (± 0.12) μg L−1, respectively. 4-NC was detected in all snowmelt outflow samples and all but one snow samples (88 % detection frequency) but was below the limit of quantification for the given method. All samples were collected during a sampling regime at the Palisade Glacier in August of 2021. Quantum yields of photolysis at the 277 nm absorption band were determined to be 0.36 (±0.06) and 0.21 (±0.06) in ultrapure water and liquid snow, respectively. Under clear sky conditions at the Palisade Glacier, half-lives for 4-NP are estimated to range from 235 to 251 h (9.8–10.5 days) based on assays conducted in liquid snowmelt and irradiance modeling. These results suggest that the photolysis of 4-NP, and hence the production of 4-NC, is occurring at significant rates within the snowpack where 4-NC is inevitably released to downstream catchment areas via snowmelt. 4-NC is significantly more toxic than its precursor, thereby raising amplified concerns for downstream human and wildlife populations. Furthermore, the ubiquity of 4-NP among the Earth's environments presents this as an issue of potentially global concern.