Hemilabile and Redox-Active Quinone Ligands Unlock sp3-Rich Couplings in Nickel-Catalyzed Olefin Carbosulfenylation.

A three-component coupling approach toward structurally complex dialkylsulfides is described via the nickel-catalyzed 1,2-carbosulfenylation of unactivated alkenes with organoboron nucleophiles and alkylsulfenamide (N-S) electrophiles. Efficient catalytic turnover is facilitated using a tailored N-S electrophile containing an N-methyl methanesulfonamide leaving group, allowing catalyst loadings as low as 1 mol%. Regioselectivity is controlled by a collection of monodentate, weakly coordinating native directing groups, including sulfonamides, amides, sulfinamides, phosphoramides, and carbamates. Key to the development of this transformation is the identification of quinones as a family of hemilabile and redox-active ligands that tune the steric and electron properties of the metal throughout the catalytic cycle. DFT calculations show that the duroquinone (DQ) ligand adopts different coordination modes in different stages of the Ni-catalyzed 1,2-carbosulfenylation-binding as an η6 capping ligand to stabilize the precatalyst/resting state and prevent catalyst decomposition, binding as an X-type redox-active durosemiquinone radical anion to promote alkene migratory insertion with a less distorted square planar Ni(II) center, while binding as an η1 L-type ligand to promote N-S oxidative addition at a relatively more electron-rich and sterically less crowded Ni(I) center.

Novel Insights into the Antimicrobial and Antibiofilm Activity of Pyrroloquinoline Quinone (PQQ); In Vitro, In Silico, and Shotgun Proteomic Studies.

Microbial infections pose a significant global health threat, affecting millions of individuals and leading to substantial mortality rates. The increasing resistance of microorganisms to conventional treatments requires the development of novel antimicrobial agents. Pyrroloquinoline quinone (PQQ), a natural medicinal drug involved in various cellular processes, holds promise as a potential antimicrobial agent. In the present study, our aim was, for the first time, to explore the antimicrobial activity of PQQ against 29 pathogenic microbes, including 13 fungal strains, 8 Gram-positive bacteria, and 8 Gram-negative bacteria. Our findings revealed potent antifungal properties of PQQ, particularly against Syncephalastrum racemosum, Talaromyces marneffei, Candida lipolytica, and Trichophyton rubrum. The MIC values varied between fungal strains, and T. marneffei exhibited a lower MIC, indicating a greater susceptibility to PQQ. In addition, PQQ exhibited notable antibacterial activity against Gram-positive and -negative bacteria, with a prominent inhibition observed against Staphylococcus epidermidis, Proteus vulgaris, and MRSA strains. Remarkably, PQQ demonstrated considerable biofilm inhibition against the MRSA, S. epidermidis, and P. vulgaris strains. Transmission electron microscopy (TEM) studies revealed that PQQ caused structural damage and disrupted cell metabolism in bacterial cells, leading to aberrant morphology, compromised cell membrane integrity, and leakage of cytoplasmic contents. These findings were further affirmed by shotgun proteomic analysis, which revealed that PQQ targets several important cellular processes in bacteria, including membrane proteins, ATP metabolic processes, DNA repair processes, metal-binding proteins, and stress response. Finally, detailed molecular modeling investigations indicated that PQQ exhibits a substantial binding affinity score for key microbial targets, including the mannoprotein Mp1P, the transcriptional regulator TcaR, and the endonuclease PvuRTs1I. Taken together, our study underscores the effectiveness of PQQ as a broad-spectrum antimicrobial agent capable of combating pathogenic fungi and bacteria, while also inhibiting biofilm formation and targeting several critical biological processes, making it a promising therapeutic option for biofilm-related infections.

Beyond adult models: Tribolium castaneum larval timekeeping reveals unexpected robustness and insights into circadian clock.

Circadian rhythms are self-sustained endogenous oscillations that are found in all living organisms. In insects, circadian rhythms control a wide variety of behavioral and physiological processes, including feeding, locomotion, mating, and metabolism. While the role of circadian rhythms in adult insects is well-understood, it is largely unexplored in larvae. This study investigates the potential for larval synchronized activity in the red flour beetle (Tribolium castaneum), a species exhibiting solitary and aggregation phases. We hypothesized that, similar to adults, larvae would exhibit a daily activity pattern governed by an endogenous circadian clock. We further predicted that the transition between the solitary and gregarious phases extends to unique temporal activity patterns. Our results revealed unique timekeeper gene expression in larvae, leading to a distinct daily rhythm characterized by nocturnal activity. Cues indicating on potential cannibalism did not change daily activity peak. However, the absence of these cues significantly reduced the proportion of rhythmic larvae and led to higher variation in peak activity, highlighting the crucial role of social interactions in shaping their rhythmicity. This study sheds light on the evolution and function of larval synchronization in group-living insects, offering novel insights into this complex behavior.

Unveiling the kinetic versatility of aryl-alcohol oxidases with different electron acceptors.

Introduction: Aryl-alcohol oxidase (AAO) shows a pronounced duality as oxidase and dehydrogenase similar to that described for other glucose-methanol-choline (GMC) oxidase/dehydrogenase superfamily proteins involved in lignocellulose decomposition. In this work, we detail the overall mechanism of AAOs from Pleurotus eryngii and Bjerkandera adusta for catalyzing the oxidation of natural aryl-alcohol substrates using either oxygen or quinones as electron acceptors and describe the crystallographic structure of AAO from B. adusta in complex with a product analogue. Methods: Kinetic studies with 4-methoxybenzyl and 3-chloro-4- methoxybenzyl alcohols, including both transient-state and steady-state analyses, along with interaction studies, provide insight into the oxidase and dehydrogenase mechanisms of these enzymes. Moreover, the resolution of the crystal structure of AAO from B. adusta allowed us to compare their overall folding and the structure of the active sites of both AAOs in relation to their activities. Results and Discussion: Although both enzymes show similar mechanistic properties, notable differences are highlighted in this study. In B. adusta, the AAO oxidase activity is limited by the reoxidation of the flavin, while in P. eryngii the slower step takes place during the reductive half-reaction, which determines the overall reaction rate. By contrast, dehydrogenase activity in both enzymes, irrespective of the alcohol participating in the reaction, is limited by the hydroquinone release from the active site. Despite these differences, both AAOs are more efficient as dehydrogenases, supporting the physiological role of this activity in lignocellulosic decay. This dual activity would allow these enzymes to adapt to different environments based on the available electron acceptors.

Elucidation and active ingredient identification of aqueous extract of Ficus exasperata Vahl leaf against bisphenol A-induced toxicity through in vivo and in silico assessments.

Bisphenol A (BPA), an endocrine-disrupting chemical, poses significant health problems due to its induction of oxidative stress, inflammation, etc. Whereas Ficus exasperata Vahl leaf (FEVL) was reported for its ethnopharmacological properties against several ailments owing to its antioxidant, anti-inflammatory properties, etc. Here, we aim to elucidate and identify the bioactive compounds of aqueous extract of FEVL (AEFEVL) against BPA-induced toxicity using in vivo and in silico assessments. To determine the BPA toxicity mechanism and safe doses of AEFEVL, graded doses of BPA (0-400 µM) and AEFEVL (0-2.0 mg/10 g diets) were separately fed to flies to evaluate survival rates and specific biochemical markers. The mitigating effect of AEFEVL (0.5 and 1.0 mg/10 g diet) against BPA (100 and 200 µM)-induced toxicity in the flies after 7-day exposure was also carried out. Additionally, molecular docking analysis of BPA and BPA-o-quinone (BPAQ) against selected antioxidant targets, and HPLC-MS-revealed AEFEVL compounds against Keap-1 and IKKß targets, followed by ADMET analysis, was conducted. Emergence rate, climbing ability, acetylcholinesterase, monoamine oxidase-B, and glutathione-S-transferase activities, and levels of total thiols, non-protein thiols, nitric oxide, protein carbonyl, malondialdehyde, and cell viability were evaluated. BPA-induced altered biochemical and behavioral parameters were significantly mitigated by AEFEVL in the flies (p < 0.05). BPAQ followed by BPA exhibited higher inhibitory activity, and epigallocatechin (EGC) showed the highest inhibitory activity among the AEFEVL compounds with desirable ADMET properties. Conclusively, our findings revealed that EGC might be responsible for the mitigative effect displayed by AEFEVL in BPA-induced toxicity in D. melanogaster.

Discovery of a potent melatonin-based inhibitor of quinone reductase-2 with neuroprotective and neurogenic properties.

5-Methoxy-3-(5-methoxyindolin-2-yl)-1H-indole (3), whose structure was unambiguously elucidated by X-ray analysis, was identified as a multi-target compound with potential application in neurodegenerative diseases. It is a low nanomolar inhibitor of QR2 (IC50 = 7.7 nM), with greater potency than melatonin and comparable efficacy to the most potent QR2 inhibitors described to date. Molecular docking studies revealed the potential binding mode of 3 to QR2, which explains its superior potency compared to melatonin. Furthermore, compound 3 inhibits hMAO-A, hMAO-B and hLOX-5 in the low micromolar range and is an excellent ROS scavenger. In phenotypic assays, compound 3 showed neuroprotective activity in a cellular model of oxidative stress damage, it was non-toxic, and was able to activate neurogenesis from neural stem-cell niches of adult mice. These excellent biological properties, together with its both good in silico and in vitro drug-like profile, highlight compound 3 as a promising drug candidate for neurodegenerative diseases.

Structure-driven development of a biomimetic rare earth artificial metalloprotein.

The 2011 discovery of the first rare earth-dependent enzyme in methylotrophic Methylobacterium extorquens AM1 prompted intensive research toward understanding the unique chemistry at play in these systems. This enzyme, an alcohol dehydrogenase (ADH), features a La3+ ion closely associated with redox-active coenzyme pyrroloquinoline quinone (PQQ) and is structurally homologous to the Ca2+-dependent ADH from the same organism. AM1 also produces a periplasmic PQQ-binding protein, PqqT, which we have now structurally characterized to 1.46-Å resolution by X-ray diffraction. This crystal structure reveals a Lys residue hydrogen-bonded to PQQ at the site analogously occupied by a Lewis acidic cation in ADH. Accordingly, we prepared K142A- and K142D-PqqT variants to assess the relevance of this site toward metal binding. Isothermal titration calorimetry experiments and titrations monitored by UV-Vis absorption and emission spectroscopies support that K142D-PqqT binds tightly (Kd = 0.6 ± 0.2 µM) to La3+ in the presence of bound PQQ and produces spectral signatures consistent with those of ADH enzymes. These spectral signatures are not observed for WT- or K142A-variants or upon addition of Ca2+ to PQQ ⸦ K142D-PqqT. Addition of benzyl alcohol to La3+-bound PQQ ⸦ K142D-PqqT (but not Ca2+-bound PQQ ⸦ K142D-PqqT, or La3+-bound PQQ ⸦ WT-PqqT) produces spectroscopic changes associated with PQQ reduction, and chemical trapping experiments reveal the production of benzaldehyde, supporting ADH activity. By creating a metal binding site that mimics native ADH enzymes, we present a rare earth-dependent artificial metalloenzyme primed for future mechanistic, biocatalytic, and biosensing applications.

Synthesis of melanin-like amino acid surfactant with enzymatic hydrolysates from silk degumming water.

The degummed wastewater from silk processing contains a huge amount of amino acids and polypeptides from sericin. The silk degumming water is far from being exploited fully. Sericin in the degumming water is generally wasted and causes environmental pollution. In this study, simulated silk degumming water was hydrolyzed by alkaline protease to produce abundant amino acids and polypeptides. After enzymatic hydrolysis, the maximum free amino groups concentration in the silk degumming water was approximately 54 mM. It facilitated the recycling of silk degumming water for the production of melanin-like amino acid surfactants as raw materials. 4-Tert-butylcatechol was used as the starting material to generate o-quinone via oxidation by ceric ammonium nitrate. o-Quinone was coupled with free amino groups in enzymatic hydrolysates of silk degumming water to synthesize a sericin-based amino acid surfactant as hydrophobic and hydrophilic group, respectively. Through the green and simple synthesis route, the product was characterized to have a novel melanin-like structure. The product exhibited superior surface-active properties by lowering the surface tension to 32.39 mN m-1. Furthermore, it demonstrated good foaming ability and foam stability, with the initial foam volume of 37 mL and the foam half-life time of more than 25 min. The product owned a good emulsification ability in the oil-water emulsion with delamination time of 297 s and 291 s for emulsion formed by soybean oil and liquid paraffin, respectively. The wetting time of the canvas sheet was only 134 s. Consequently, the product showed low surface tension, good foaming, emulsifying, and wetting properties.

Regulation of folate transport at the mouse arachnoid barrier.

BACKGROUND: Folates are a family of B9 vitamins essential for normal growth and development in the central nervous system (CNS). Transport of folates is mediated by three major transport proteins: folate receptor alpha (FRα), proton-coupled folate transporter (PCFT), and reduced folate carrier (RFC). Brain folate uptake occurs at the choroid plexus (CP) epithelium through coordinated actions of FRα and PCFT, or directly into brain parenchyma at the vascular blood-brain barrier (BBB), mediated by RFC. Impaired folate transport can occur due to loss of function mutations in FRα or PCFT, resulting in suboptimal CSF folate levels. Our previous reports have demonstrated RFC upregulation by nuclear respiratory factor-1 (NRF-1) once activated by the natural compound pyrroloquinoline quinone (PQQ). More recently, we have identified folate transporter localization at the arachnoid barrier (AB). The purpose of the present study was to further characterize folate transporters localization and function in AB cells, as well as their regulation by NRF-1/PGC-1α signaling and folate deficiency. METHODS: In immortalized mouse AB cells, polarized localization of RFC and PCFT was assessed by immunocytochemical analysis, with RFC and PCFT functionality examined with transport assays. The effects of PQQ treatment on changes in RFC functional expression were also investigated. Mouse AB cells grown in folate-deficient conditions were assessed for changes in gene expression of the folate transporters, and other key transporters and tight junction proteins. RESULTS: Immunocytochemical analysis revealed apical localization of RFC at the mouse AB epithelium, with PCFT localized on the basolateral side and within intracellular compartments. PQQ led to significant increases in RFC functional expression, mediated by activation of the NRF-1/PGC-1α signalling cascade. Folate deficiency led to significant increases in expression of RFC, MRP3, P-gp, GLUT1 and the tight junction protein claudin-5. CONCLUSION: These results uncover the polarized expression of RFC and PCFT at the AB, with induction of RFC functional expression by activation of the NRF-1/PGC-1α signalling pathway and folate deficiency. These results suggest that the AB may contribute to the flow of folates into the CSF, representing an additional pathway when folate transport at the CP is impaired.

Correlation of Vanillin-Induced Cytotoxicity with CYP3A-Mediated Metabolic Activation.

Vanillin (VAN) is a common flavoring agent that can cause liver damage when ingested in large amounts. Nevertheless, the precise processes responsible for its toxicity remain obscure. The present research aimed to examine the metabolic activation of VAN and establish a potential correlation between its reactive metabolites and its cytotoxicity. In rat liver microsomes incubated with VAN, reduced glutathione/N-acetylcysteine (GSH/NAC), and nicotinamide adenine dinucleotide phosphate (NADPH), two conjugates formed from GSH and one conjugate derived from NAC were identified. We also discovered one GSH conjugate in both the bile obtained from rats and the rat primary hepatocytes that were subjected to VAN exposure. Additionally, the NAC conjugate exerted in the urine of VAN-treated rats was observed. These results indicate that a quinone intermediate was produced from VAN both in vitro and in vivo. Next, we identified CYP3A as the main enzyme that initiated the bioactive pathway of VAN. After the activity of CYP3A was selectively inhibited by ketoconazole (KTZ), the generation of the GSH conjugate declined in hepatocytes exposed to VAN. Furthermore, the vulnerability to VAN-induced toxicity was alleviated by KTZ in hepatocytes. Thus, we propose that the cytotoxicity of VAN may derive from metabolic activation triggered by CYP3A.

Cytotoxic and Antitumor Agents from Genus Rubia.

Genus Rubia is widely distributed in almost all regions of the world, with 36 species and 2 varieties in China. Rubia species, such as Rubia cordifolia, have been used in traditional Chinese medicine for the treatment of diseases since ancient times. In recent years, the study of anticancer effects in traditional Chinese medicine has become a popular topic, and some studies have shown that several Rubia species extracts have cytotoxic and antitumor effects, and some of them have been shown to contain specific antitumor agents. Therefore, this review focuses on the cytotoxic and antitumor effects of the chemical constituents contained in Genus Rubia. Summarized 71 types of chemical substances in 5 categories with the effect of cytotoxicity and antitumor, as well as their structures, targets and mechanisms of action.

Unveiling the potential: exploring the efficacy of complex III inhibitors in fungal disease control.

Phytopathogenic fungi are a key challenge to maximizing crop yield and quality for a growing global population. In this review, we give an overview of representative compounds that inhibit complex III, also known as bc1 complex, covering quinone inside inhibitors, quinone outside inhibitors, and quinone inside and outside inhibitors via the stigmatellin binding mode. Novel solutions to the escalating problem of resistance are still required, therefore compounds with alternative scaffolds, alternative docking modes, different mechanisms of action and improved efficacy against complex III necessitate ongoing research. © 2024 Society of Chemical Industry.

Oxidative [4+2] Annulation of Pyrrole-2-carbaldehyde Derivatives with o-Hydroxyphenyl Propargylamines: Syntheses of 5,6,7-Trisubstituted Indolizines.

A base promoted oxidative [4+2] annulation of pyrrole-2-carbaldehyde derivatives with o-hydroxyphenyl propargylamines for the synthesis of highly substituted indolizines has been developed. Using DBN as base, a broad range of 5,6,7-trisubstituted indolizines have been prepared in good to excellent yields under mild conditions, and many useful functional groups can be tolerated.

Smart RNA Sequencing Reveals the Toxicological Effects of Diisobutyl Phthalate (DiBP) in Porcine Oocytes.

Diisobutyl phthalate (DiBP) is commonly used in the plastics industry, and recent studies have shown that environmental exposure and accumulation in the food chain caused inflammation in some organs. However, the underlying mechanisms by which DiBP affects oocyte quality have not yet been fully defined. We used immunostaining and fluorescence to evaluate the effects of DiBP exposure and demonstrated that it impaired the morphology of matured porcine oocytes through generation of cytoplasmic fragmentation, accompanied by the perturbed dynamics of the spindle and actin cytoskeleton, misdistributed endoplasmic reticulum, as well as partial exocytosis of cortical granules and ovastacin. Moreover, analysis of Smart RNA-seq found that DiBP-induced aberrant oocyte maturation could be induced by abnormal mitochondrial function and apoptosis. Importantly, we discovered that supplementation with pyrroloquinoline quinone (PQQ) significantly attenuated the meiotic abnormalities induced by DiBP exposure through the modulation of reactive oxygen species levels. Our findings demonstrated that DiBP exposure adversely affects oocyte meiotic maturation and that PQQ supplementation was an effective strategy to protect oocyte quality against DiBP exposure.

Lewis Acid-Catalyzed Unusual (4+3) Annulation of para-Quinone Methides with Bicyclobutanes: Access to Oxabicyclo[4.1.1] octanes.

Over the past few years, there has been a surge of interest in the chemistry of bicyclobutanes (BCBs). Although BCBs have been used to synthesize bicyclo[2.1.1]hexanes and bicyclo[3.1.1]heptanes, the synthesis of bicyclo[4.1.1]octanes has remained elusive. Herein, we report the first Lewis acid-catalyzed unexpected (4+3) annulation of para-quinonemethides (p-QMs) with BCBs allowing the synthesis of oxabicyclo[4.1.1]octanes proceeding under mild conditions. With 5 mol % of Bi(OTf)3, the reaction afforded the (4+3) annulated product in high regioselectivity and good functional group compatibility via a simultaneous Lewis acid activation of BCBs and p-QMs. The reaction is likely initiated by the 1,6-addition of Lewis acid activated BCBs to p-QMs followed by the C2-selective intramolecular addition of the phenol moiety to the generated cyclobutyl cation intermediate. Moreover, detailed mechanistic studies provided insight into the mechanism of the reaction.

Screening and identification of peptidyl arginine deiminase 4 inhibitors from herbal plants extracts and purified natural products by a trypsin assisted sensitive immunoassay based on streptavidin magnetic beads.

Peptidyl arginine deiminase 4 (PAD4) plays a critical role in many autoimmune diseases including rheumatoid arthritis. Herein, a trypsin assisted highly immunoassay method was established to determine PAD4 activity and screen potent inhibitors from herbal plants extracts and purified natural products. The method was applied to determine endogenous PAD4 activity in both cell and tissue lysates, as well as the inhibitory effects of 20 herbal plants and 50 purified natural products. The Cinnamomi ramulus extract showed strongest inhibitory potency with IC50 value lower than 5 µg/mL. Meanwhile, pyrroloquinoline quinone (PQQ), widely used as a dietary supplement, was discovered as a promising PAD4 inhibitor with an IC50 value lower than 4 µM. The inhibition kinetic analysis, drug affinity response target stability (DARTS) and molecular docking were performed to confirm the interaction between PQQ and PAD4. This method has great potential for researchers to monitor activities and discover potential inhibitors of PAD4.

Reactions between ferric oxyhydroxide mineral coatings and a dimethoxyhydroquinone: A source of hydroxyl radicals.

Quinones are organic molecules that facilitate electron-transfer reactions in terrestrial environments. The reduced forms, hydroquinones, are powerful reductants that can trigger non-enzymatic radical-based decomposition of organic matter and contaminants by simultaneous reduction of iron and oxygen. Iron oxides often occur as coatings on other minerals, thus our study investigated the reactions between the ferric oxyhydroxide (FeO(OH)) surface coatings on gibbsite (Al(OH)3) and 2,6-dimethoxy-1,4-hydroquinone (2,6-DMHQ). The main aim was to investigate the oxidation of 2,6-DMHQ and the generation ∙OH in the presence of O2 at low Fe concentrations in a novel setup that allows local structural characterization. The heterogeneous redox reactions between 2,6-DMHQ and the FeO(OH) coatings were studied at pH 5.0 as a function of the amount of Fe present on the gibbsite surfaces, including the effect of aging of the FeO(OH) coatings. The results showed that reactions between 2,6-DMHQ and FeO(OH) coated gibbsite under ambient conditions can generate substantial amounts of ·OH, comparable with amounts generated on pure ferrihydrite surfaces. The ·OH is the product of two sequential reactions: hydroquinone oxidation by O2 and degradation of the formed H2O2. The calculated rate constant of the former reaction is the same regardless of amount of FeO(OH) coating suggesting a surface catalytic process where 2,6-DMHQ is oxidized by O2 resulting in formation of H2O2. Subsequently, the observed induction period, the low Fe2+ (aq) concentrations in solution and the dependency of FeO(OH) coating amount influencing ·OH formation suggest that the pathway for ∙OH is through H2O2 decomposition by the surface sites on the FeO(OH) coating. Overall, this study shows that co-existence of oxygen, FeO(OH) and organic reductants, possibly secreted by soil microorganisms, creates favorable conditions for generation of ·OH contributing to decomposition of organic matter and organic pollutants in soil environments.

Alkoxyalkylation of Electron-Rich Aromatic Compounds.

Alkoxyalkylation and hydroxyalkylation methods utilizing oxo-compound derivatives such as aldehydes, acetals or acetylenes and various alcohols or water are widely used tools in preparative organic chemistry to synthesize bioactive compounds, biosensors, supramolecular compounds and petrochemicals. The syntheses of such molecules of broad relevance are facilitated by acid, base or heterogenous catalysis. However, degradation of the N-analogous Mannich bases are reported to yield alkoxyalkyl derivatives via the retro-Mannich reaction. The mutual derivative of all mentioned species are quinone methides, which are reported to form under both alkoxy- and aminoalkylative conditions and via the degradation of the Mannich-products. The aim of this review is to summarize the alkoxyalkylation (most commonly alkoxymethylation) of electron-rich arenes sorted by the methods of alkoxyalkylation (direct or via retro-Mannich reaction) and the substrate arenes, such as phenolic and derived carbocycles, heterocycles and the widely examined indole derivatives.

Effects of environmental concentrations of 6PPD and its quinone metabolite on the growth and reproduction of freshwater cladoceran.

The widespread occurrence and accumulation of N-(1,3-dimethylbutyl)-N'-phenyl-p-phenylenediamine (6PPD) and its quinone metabolite, 6PPD quinone (6PPD-Q), have been globally recognized as a critical environmental issue. However, knowledge on the adverse effects of 6PPD and 6PPD-Q on freshwater invertebrates is limited. This study investigated the effects of 6PPD and its oxidative byproduct, 6PPD-Q, on the growth and reproduction of Daphnia pulex. Through 21-day exposure experiments, we measured the uptake of 0.1, 1, and 10 µg/L 6PPD and 6PPD-Q by D. pulex and assessed the effects on growth and fecundity of D. pulex. While 6PPD and 6PPD-Q did not affect the mortality rate of D. pulex, 6PPD-Q exposure inhibited the growth of D. pulex, indicating potential ecological risks. In particular, the reproductive capacity of D. pulex remained unaffected across the tested concentrations of 6PPD and 6PPD-Q, suggesting specific toxicological pathways that warrant further investigation. This study underscored the importance of evaluating the sublethal effects of emerging contaminants such as 6PPD and 6PPD-Q on aquatic invertebrates, and highlighted the need for comprehensive risk assessments to better understand their environmental impacts.

Corynebacterium glutamicum pyruvate:quinone oxidoreductase: an enigmatic metabolic enzyme with unusual structural features.

Pyruvate:quinone oxidoreductase (PQO) is a flavin-containing peripheral membrane enzyme catalyzing the decarboxylation of pyruvate to acetate and CO2 with quinone as an electron acceptor. Here, we investigate PQO activity in Corynebacterium glutamicum, examine purified PQO, and describe the crystal structure of the native enzyme and a truncated version. The specific PQO activity was highest in stationary phase cells grown in complex medium, lower in cells grown in complex medium containing glucose or acetate, and lowest in cells grown in minimal acetate-medium. A similar pattern with about 30-fold higher specific PQO activities was observed in C. glutamicum with plasmid-bound pqo expression under the control of the tac promoter, indicating that the differences in PQO activity are likely due to post-transcriptional control. Continuous cultivation of C. glutamicum at dilution rates between 0.05 and 0.4 h-1 revealed a negative correlation between PQO activity and growth rate. Kinetic analysis of PQO enzymes purified from cells grown in complex or in minimal acetate-medium revealed substantial differences in specific activity (72.3 vs. 11.9 U·mg protein-1) and turnover number (kcat: 440 vs. 78 s-1, respectively), suggesting post-translational modifications affecting PQO activity. Structural analysis of PQO revealed a homotetrameric arrangement very similar to the Escherichia coli pyruvate oxidase PoxB except for the C-terminal membrane binding domain, which exhibited a conformation markedly different from its PoxB counterpart. A truncated PQO variant lacking 17 C-terminal amino acids showed higher affinity to pyruvate and was independent of detergent activation, highlighting the importance of the C-terminus for enzyme activation and lipid binding.