Identification of a novel xanthine oxidoreductase inhibitor for hyperuricemia treatment with high efficacy and safety profile.

Hyperuricemia is with growing incidence and of high risk to develop into gout and other metabolic diseases. The key enzyme catalyzing uric acid synthesis, xanthine oxidoreductase (XOR) is a vital target for anti-hyperuricemic drugs, while XOR inhibitors characterized as both potent and safe are currently in urgent need. In this study, a novel small molecule compound, CC15009, was identified as a specific XOR inhibitor. CC15009 exerted strongest in vitro XOR inhibitory activity among current XOR inhibitors. It also showed favorable dose-dependent uric acid-lowering effects in two different XOR substrate-induced hyperuricemic mouse models, which was significantly superior than the current first-line drug, allopurinol. Mechanically, the direct binding of CC15009 against XOR was confirmed by molecular docking and SPR analysis. The inhibition mode was competitive and reversible. Besides, the potential antioxidant activity of CC15009 was indicated by its strong inhibitory activity against the oxidized isoform of XOR, which reduced ROS generation as the byproduct. Regarding the safety concerns of current XOR inhibitors, especially in cardiovascular risks, the safety of CC15009 was comprehensively evaluated. No significant abnormality was observed in the acute, subacute toxicity tests and mini-AMES test. Notably, there was no obvious inhibition of CC15009 against cardiac ion channels, including hERG, Nav1.5, Cav1.2 at the concentration of 30 µM, indicating its lower cardiovascular risk. Taken together, our results supported CC15009 as a candidate of high efficacy and safety profile to treat hyperuricemia through direct XOR inhibition.

Biosynthetic Strategies of Berberine Bridge Enzyme-like Flavoprotein Oxidases toward Structural Diversification in Natural Product Biosynthesis.

Berberine bridge enzyme-like oxidases are often involved in natural product biosynthesis and are seen as essential enzymes for the generation of intricate pharmacophores. These oxidases have the ability to transfer a hydride atom to the FAD cofactor, which enables complex substrate modifications and rearrangements including (intramolecular) cyclizations, carbon-carbon bond formations, and nucleophilic additions. Despite the diverse range of activities, the mechanistic details of these reactions often remain incompletely understood. In this Review, we delve into the complexity that BBE-like oxidases from bacteria, fungal, and plant origins exhibit by providing an overview of the shared catalytic features and emphasizing the different reactivities. We propose four generalized modes of action by which BBE-like oxidases enable the synthesis of natural products, ranging from the classic alcohol oxidation reactions to less common amine and amide oxidation reactions. Exploring the mechanisms utilized by nature to produce its vast array of natural products is a subject of considerable interest and can lead to the discovery of unique biochemical activities.

Heat-stress memory enhances the acclimation of a migratory insect pest to global warming.

In the face of rising global temperatures, the mechanisms behind an organism's ability to acclimate to heat stress remain enigmatic. The rice leaf folder, Cnaphalocrocis medinalis, traditionally viewed as temperature-sensitive, paradoxically exhibits robust larval acclimation to heat stress. This study used the heat-acclimated strain HA39, developed through multigenerational exposure to 39°C during the larval stage, and the unacclimated strain HA27 reared at 27°C to unravel the transgenerational effects of heat acclimation and its regulatory mechanisms. Heat acclimation for larvae incurred a fitness cost in pupae when exposed to high temperature, yet a significant transgenerational effect surfaced, revealing heightened fitness benefit in pupae from HA39, even without additional heat exposure during larval recovery at 27°C. This transgenerational effect exhibited a short-term memory, diminishing after two recovery generations. Moreover, the effect correlated with increased superoxide dismutase (SOD) enzyme activity and expression levels of oxidoreductase genes, representing physiological and molecular foundations of heat acclimation. Heat-acclimated larvae displayed elevated DNA methylation levels, while pupae from HA39, in recovery generations, exhibited decreased methylation indicated by the upregulation of a demethylase gene and downregulation of two methyltransferase genes at high temperatures. In summary, heat acclimation induces DNA methylation, orchestrating heat-stress memory and influencing the expression levels of oxidoreductase genes and SOD activity. Heat-stress memory enhances the acclimation of the migratory insect pest to global warming.

Structure-Activity Relationships and Changes in the Inhibition of Xanthine Oxidase by Polyphenols: A Review.

Hyperuricemia (HUA), or elevated uric acid in the blood, has become more prevalent in recent years. Polyphenols, which are known to have good inhibitory activity on xanthine oxidoreductase (XOR), are effective in uric acid reduction. In this review, we address the structure-activity relationship of flavonoids that inhibit XOR activity from two perspectives: the key residues of XOR and the structural properties of flavonoids. Flavonoids' inhibitory effect is enhanced by their hydroxyl, methoxy, and planar structures, whereas glycosylation dramatically reduces their activity. The flavonoid structure-activity relationship informed subsequent discussions of the changes that occur in polyphenols' XOR inhibitory activity during their extraction, processing, gastrointestinal digestion, absorption, and interactions. Furthermore, gastrointestinal digestion and heat treatment during processing can boost the inhibition of XOR. Polyphenols with comparable structures may have a synergistic effect, and their synergy with allopurinol thus provides a promising future research direction.

Molecular insight into the potential functional role of pseudoenzyme GFOD1 via interaction with NKIRAS2.

The glucose-fructose oxidoreductase/inositol dehydrogenase/rhizopine catabolism protein (Gfo/Idh/MocA) family includes a variety of oxidoreductases with a wide range of substrates that utilize NAD or NADP as redox cofactor. Human contains two members of this family, namely glucose-fructose oxidoreductase domain-containing protein 1 and 2 (GFOD1 and GFOD2). While GFOD1 exhibits low tissue specificity, it is notably expressed in the brain, potentially linked to psychiatric disorders and severe diseases. Nevertheless, the specific function, cofactor preference, and enzymatic activity of GFOD1 remain largely unknown. In this work, we find that GFOD1 does not bind to either NAD or NADP. Crystal structure analysis unveils that GFOD1 exists as a typical homodimer resembling other family members, but lacks essential residues required for cofactor binding, suggesting that it may function as a pseudoenzyme. Exploration of GFOD1-interacting partners in proteomic database identifies NK-κB inhibitor-interacting Ras-like 2 (NKIRAS2) as one potential candidate. Co-immunoprecipitation (co-IP) analysis indicates that GFOD1 interacts with both GTP- and GDP-bound forms of NKIRAS2. The predicted structural model of the GFOD1-NKIRAS2 complex is validated in cells using point mutants and shows that GFOD1 selectively recognizes the interswitch region of NKIRAS2. These findings reveal the distinct structural properties of GFOD1 and shed light on its potential functional role in cellular processes.

Sex-dependent differences in macaque brain mitochondria.

Mitochondrial bioenergetics in females and males is different. However, whether mitochondria from male and female brains display differences in enzymes of oxidative phosphorylation remains unknown. Therefore, we characterized mitochondrial complexes from the brains of male and female macaques (Macaca mulatta). Cerebral tissue from male macaques exhibits elevated content and activity of mitochondrial complex I (NADH:ubiquinone oxidoreductase) and higher activity of complex II (succinate dehydrogenase) compared to females. No significant differences between sexes were found in the content of α-ketoglutarate dehydrogenase or in the activities of cytochrome c oxidase and F1Fo ATPase. Our results underscore the need for further investigations to elucidate sex-related mitochondrial differences in humans.

Glycoside-metabolizing oxidoreductase D3dgpA from human gut bacterium.

The Gfo/Idh/MocA family enzyme DgpA was known to catalyze the regiospecific oxidation of puerarin to 3"-oxo-puerarin in the presence of 3-oxo-glucose. Here, we discovered that D3dgpA, dgpA cloned from the human gut bacterium Dorea sp. MRG-IFC3, catalyzed the regiospecific oxidation of various C-/O-glycosides, including puerarin, in the presence of methyl ß-D-3-oxo-glucopyranoside. While C-glycosides were converted to 3"- and 2"-oxo-products by D3dgpA, O-glycosides resulted in the formation of aglycones and hexose enediolone from the 3"-oxo-products. From DFT calculations, it was found that isomerization of 3"-oxo-puerarin to 2"-oxo-puerarin required a small activation energy of 9.86 kcal/mol, and the O-glycosidic bond cleavage of 3"-oxo-products was also thermodynamically favored with a small activation energy of 3.49 kcal/mol. In addition, the reaction mechanism of D3dgpA was discussed in comparison to those of Gfo/Idh/MocA and GMC family enzymes. The robust reactivity of D3dgpA was proposed as a new general route for derivatization of glycosides.

The Cellular Genesis of Metabolic Syndrome and the Role of Anti-urate Drugs in Hyperuricemia Patients: A Systematic Review.

Hyperuricemia results due to the underexcretion of uric acid through kidneys or overproduction due to either intake of purine-rich foods, a high caloric diet, or a decreased activity of purine recycler hypoxanthine-guanine phosphoribosyl transferase (HGPRT). Increased xanthine oxidoreductase (XOR) enzyme activity may contribute to hyperuricemia. Literature provides growing evidence that an independent component that contributes to the development of metabolic syndrome (MetS) and associated comorbidities is hyperuricemia. Thus, precise cellular mechanisms involved during MetS and related comorbidities in hyperuricemia, and the role of anti-urate medicines in these mechanisms require further investigations. We searched online libraries PubMed and Google Scholar for data collection. We used Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) 2020 guidelines for literature identification, selection, screening, and determining eligibility to produce unbiased meaningful outcomes. We applied quality assessment tools for the quality appraisal of the studies. And, outcomes were extracted from the selected studies, which revealed the relationship between hyperuricemia and MetS components by causing inflammation, endothelial dysfunction, oxidative stress, and endoplasmic reticulum stress. The selected studies reflected the role of xanthine oxide (XO) inhibitors beyond inhibition. This systematic review concluded that hyperuricemia independently causes inflammation, oxidative stress, endothelial damage, and endoplasmic reticulum stress in patients with hyperuricemia. These mechanisms provide a cellular basis for metabolic syndrome and related comorbidities. In this context, XO inhibitors and their beneficial effects go beyond XOR inhibition to ameliorate these pathological mechanisms.

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.

Rv0547c, a functional oxidoreductase, supports Mycobacterium tuberculosis persistence by reprogramming host mitochondrial fatty acid metabolism.

Mycobacterium tuberculosis (Mtb) successfully thrives in the host by adjusting its metabolism and manipulating the host environment. In this study, we investigated the role of Rv0547c, a protein that carries mitochondria-targeting sequence (MTS), in mycobacterial persistence. We show that Rv0547c is a functional oxidoreductase that targets host-cell mitochondria. Interestingly, the localization of Rv0547c to mitochondria was independent of the predicted MTS but depended on specific arginine residues at the N- and C-terminals. As compared to the mitochondria-localization defective mutant, Rv0547c-2SDM, wild-type Rv0547c increased mitochondrial membrane fluidity and spare respiratory capacity. To comprehend the possible reason, comparative lipidomics was performed that revealed a reduced variability of long-chain and very long-chain fatty acids as well as altered levels of phosphatidylcholine and phosphatidylinositol class of lipids upon expression of Rv0547c, explaining the increased membrane fluidity. Additionally, the over representation of propionate metabolism and ß-oxidation intermediates in Rv0547c-targeted mitochondrial fractions indicated altered fatty acid metabolism, which corroborated with changes in oxygen consumption rate (OCR) upon etomoxir treatment in HEK293T cells transiently expressing Rv0547c, resulting in enhanced mitochondrial fatty acid oxidation capacity. Furthermore, Mycobacterium smegmatis over expressing Rv0547c showed increased persistence during infection of THP-1 macrophages, which correlated with its increased expression in Mtb during oxidative and nutrient starvation stresses. This study identified for the first time an Mtb protein that alters mitochondrial metabolism and aids in survival in host macrophages by altering fatty acid metabolism to its benefit and, at the same time increases mitochondrial spare respiratory capacity to mitigate infection stresses and maintain cell viability.

Association Between NAD(P)H Quinone Oxidoreductase 1 (NQO1) Gene Methylation/Expression and Bleeding Incidence Among an Iranian Population Undergoing Warfarin Therapy.

Increased bleeding tendency is a common and challenging complication of warfarin therapy which results in extensive pharmacogenomic studies in order to develop a personalized dosing approach and minimize the risk of related side effects. Here we aimed to explore the potential role of NQO1 gene expression in warfarin response in a group of Iranian patients. We also evaluated the NQO1 promoter methylation and its association with mRNA expression. A total of 87 patients on warfarin therapy including 34 cases with drug-induced bleeding events and 53 matched controls without bleedings were included in the study. The expression of NQO1 was examined by real-time q-PCR and the methylation status of its promoter region was analyzed using methyQESD technique. There was a significant association between the reduced NQO1 gene expression and susceptibility to bleeding before (OR = 1.92, 95% CI = 1.23-3.00, p = 0.004) and following adjustment for hypertriglyceridemia (OR = 2.22, 95% CI = 1.33-3.69, p = 0.002). Furthermore, a medium negative correlation was observed between NQO1 expression and its promoter methylation (r = - 0.382, p = 0.001). The lower expression of NQO1 which partly arises from increased methylation of promoter region, may predispose warfarin treated patients to bleeding events.

Targeting Liver Xor by GalNAc-siRNA Is an Effective Strategy for Hyperuricemia Therapy.

Hyperuricemia, i.e., increased plasma uric acid concentration, is a common problem in clinical practice, leading to gout or nephrolithiasis, and is associated with other disorders, such as metabolic syndrome, cardiovascular disease, and chronic renal disease. Xanthine oxidoreductase (XOR) is a critical rate-limiting enzyme involved in uric acid synthesis and a promising target for hyperuricemia therapy. However, XOR inhibitors currently face clinical problems such as a short half-life and side effects. Here, we found that specifically targeting liver Xor with GalNAc-siRNAs had a good therapeutic effect on hyperuricemia. First, siRNAs were designed to target various sites in the homologous region between Homo sapiens and Mus musculus Xor mRNA and were screened in primary mouse hepatocytes. Then, the siRNAs were modified to increase their stability in vivo and conjugated with GalNAc for liver-specific delivery. The effects of GalNAc-siRNAs were evaluated in three hyperuricemia mouse models, including potassium oxonate and hypoxanthine administration in WT and humanized XDH mice and Uox knockout mice. Febuxostat, a specific XOR inhibitor used for hyperuricemia treatment, was used as a positive control. Targeting liver Xor with GalNAc-siRNAs by subcutaneous administration reduced plasma uric acid levels, uric acid accumulation in the kidney, renal inflammation, and fibrosis, thereby alleviating kidney damage in hyperuricemia mouse models without hepatoxicity. The results demonstrated that targeting liver Xor with GalNAc-siRNAs was a promising strategy for hyperuricemia therapy.

Study on Oleum cinnamomi Inhibiting Cutibacterium acnes and Its Covalent Inhibition Mechanism.

Oleum cinnamomi (OCM) is a volatile component of the Cinnamomum cassia Presl in the Lauraceae family, which displays broad-spectrum antibacterial properties. It has been found that OCM has a significant inhibitory effect against Cutibacterium acnes (C. acnes), but the precise target and molecular mechanism are still not fully understood. In this study, the antibacterial activity of OCM against C. acnes and its potential effect on cell membranes were elucidated. Metabolomics methods were used to reveal metabolic pathways, and proteomics was used to explore the targets of OCM inhibiting C. acnes. The yield of the OCM was 3.3% (w/w). A total of 19 compounds were identified, representing 96.213% of the total OCM composition, with the major constituents being phenylpropanoids (36.84%), sesquiterpenoids (26.32%), and monoterpenoids (15.79%). The main component identified was trans-cinnamaldehyde (85.308%). The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of OCM on C. acnes were 60 µg/mL and 180 µg/mL, respectively. The modified proteomics results indicate that cinnamaldehyde was the main bioactive ingredient within OCM, which covalently modifies the ABC transporter adenosine triphosphate (ATP)-binding protein and nicotinamide adenine dinucleotide (NADH)-quinone oxidoreductase, hindering the amino acid transport process, and disrupting the balance between NADH and nicotinamide adenine dinucleoside phosphorus (NAD+), thereby hindering energy metabolism. We have reported for the first time that OCM exerts an antibacterial effect by covalent binding of cinnamaldehyde to target proteins, providing potential and interesting targets to explore new control strategies for gram-positive anaerobic bacteria.

Clostridioides difficile superoxide reductase mitigates oxygen sensitivity.

Clostridioides difficile causes a serious diarrheal disease and is a common healthcare-associated bacterial pathogen. Although it has a major impact on human health, the mechanistic details of C. difficile intestinal colonization remain undefined. C. difficile is highly sensitive to oxygen and requires anaerobic conditions for in vitro growth. However, the mammalian gut is not devoid of oxygen, and C. difficile tolerates moderate oxidative stress in vivo. The C. difficile genome encodes several antioxidant proteins, including a predicted superoxide reductase (SOR) that is upregulated upon exposure to antimicrobial peptides. The goal of this study was to establish SOR enzymatic activity and assess its role in protecting C. difficile against oxygen exposure. Insertional inactivation of sor rendered C. difficile more sensitive to superoxide, indicating that SOR contributes to antioxidant defense. Heterologous C. difficile sor expression in Escherichia coli conferred protection against superoxide-dependent growth inhibition, and the corresponding cell lysates showed superoxide scavenging activity. Finally, a C. difficile SOR mutant exhibited global proteome changes under oxygen stress when compared to the parent strain. Collectively, our data establish the enzymatic activity of C. difficile SOR, confirm its role in protection against oxidative stress, and demonstrate SOR's broader impacts on the C. difficile vegetative cell proteome.IMPORTANCEClostridioides difficile is an important pathogen strongly associated with healthcare settings and capable of causing severe diarrheal disease. While considered a strict anaerobe in vitro, C. difficile has been shown to tolerate low levels of oxygen in the mammalian host. Among other well-characterized antioxidant proteins, the C. difficile genome encodes a predicted superoxide reductase (SOR), an understudied component of antioxidant defense in pathogens. The significance of the research reported herein is the characterization of SOR's enzymatic activity, including confirmation of its role in protecting C. difficile against oxidative stress. This furthers our understanding of C. difficile pathogenesis and presents a potential new avenue for targeted therapies.

Redox-active vitamin C suppresses human osteosarcoma growth by triggering intracellular ROS-iron-calcium signaling crosstalk and mitochondrial dysfunction.

Pharmacological vitamin C (VC) has gained attention for its pro-oxidant characteristics and selective ability to induce cancer cell death. However, defining its role in cancer has been challenging due to its complex redox properties. In this study, using a human osteosarcoma (OS) model, we show that the redox-active property of VC is critical for inducing non-apoptotic cancer cell death via intracellular reactive oxygen species (ROS)-iron-calcium crosstalk and mitochondrial dysfunction. In both 2D and 3D OS cell culture models, only the oxidizable form of VC demonstrated potent dose-dependent cytotoxicity, while non-oxidizable and oxidized VC derivatives had minimal effects. Live-cell imaging showed that only oxidizable VC caused a surge in cytotoxic ROS, dependent on iron rather than copper. Inhibitors of ferroptosis, a form of iron-dependent cell death, along with classical apoptosis inhibitors, were unable to completely counteract the cytotoxic effects induced by VC. Further pharmacological and genetic inhibition analyses showed that VC triggers calcium release through inositol 1,4,5-trisphosphate receptors (IP3Rs), leading to mitochondrial ROS production and eventual cell death. RNA sequencing revealed down-regulation of genes involved in the mitochondrial electron transport chain and oxidative phosphorylation upon pharmacological VC treatment. Consistently, high-dose VC reduced mitochondrial membrane potential, oxidative phosphorylation, and ATP levels, with ATP reconstitution rescuing VC-induced cytotoxicity. In vivo OS xenograft studies demonstrated reduced tumor growth with high-dose VC administration, concomitant with the altered expression of mitochondrial ATP synthase (MT-ATP). These findings emphasize VC's potential clinical utility in osteosarcoma treatment by inducing mitochondrial metabolic dysfunction through a vicious intracellular ROS-iron-calcium cycle.

Construction of a Zygosaccharomyces rouxii strain overexpressing the QOR gene for increased HDMF production.

4-Hydroxy-2,5-dimethyl-3(2H)-furanone (HDMF) is a flavor compound widely found in natural products and is used in food as a flavor-enhancing agent. Quinone oxidoreductase (QOR) was verified as a key enzyme to synthesize HDMF in strawberry, while its impact on HDMF production by Zygosaccharomyces rouxii was still unknown. The QOR gene was cloned and overexpressed in Z. rouxii, and its impact on HDMF production by Z. rouxii was then further analyzed. At the same time, it is expected to obtain engineered strains of Z. rouxii with high HDMF production. The results showed that the engineered strains of Z. rouxii exhibit different levels of QOR gene expression and HDMF production; among them, the QOR6 strain exhibiting the highest gene expression level and HDMF production was named as ZrQOR. The HDMF production of the ZrQOR strain was significantly higher than that of wild-type Z. rouxii at 3 and 5 days of culture, with 1.41-fold and 1.08-fold increases, respectively. At 3 days of fermentation, the highest HDMF yield of ZrQOR strain was obtained (2.75 mg/L), 2 days ahead of the reported highest HDMF production by Z. rouxii. At 3, 5, and 7 days, QOR gene expression was 4.8-fold, 3.3-fold, and 5.6-fold higher in the ZrQOR strain than in the wild-type Z. rouxii, respectively. Therefore, overexpression of the QOR gene facilitates HDMF synthesis. The genetic stability of the 0-20 generation ZrQOR strain was stable, and there was no significant difference in colony shape, QOR expression, or HDMF production compared to the wild type. In this study, the genetic engineering Z. rouxii strain was used to improve HDMF production. This research has laid the groundwork for further industrial production of HDMF via microbial synthesis.

Hydrogen sulfide maintains mitochondrial homeostasis and regulates ganoderic acids biosynthesis by SQR under heat stress in Ganoderma lucidum.

Hydrogen sulfide (H2S) has recently been recognized as an important gaseous transmitter with multiple physiological effects in various species. Previous studies have shown that H2S alleviated heat-induced ganoderic acids (GAs) biosynthesis, an important quality index of Ganoderma lucidum. However, a comprehensive understanding of the physiological effects and molecular mechanisms of H2S in G. lucidum remains unexplored. In this study, we found that heat treatment reduced the mitochondrial membrane potential (MMP) and mitochondrial DNA copy number (mtDNAcn) in G. lucidum. Increasing the intracellular H2S concentration through pharmacological and genetic means increased the MMP level, mtDNAcn, oxygen consumption rate level and ATP content under heat treatment, suggesting a role for H2S in mitigating heat-caused mitochondrial damage in G. lucidum. Further results indicated that H2S activates sulfide-quinone oxidoreductase (SQR) and complex III (Com III), thereby maintaining mitochondrial homeostasis under heat stress in G. lucidum. Moreover, SQR also mediated the negative regulation of H2S to GAs biosynthesis under heat stress. Furthermore, SQR might be persulfidated under heat stress in G. lucidum. Thus, our study reveals a novel physiological function and molecular mechanism of H2S signalling under heat stress in G. lucidum with broad implications for research on the environmental response of microorganisms.

Dimerization of assimilatory NADPH-dependent sulfite reductase reveals elements for diflavin reductase binding at a minimal interface.

Escherichia coli NADPH-dependent assimilatory sulfite reductase is responsible for fixing sulfur for incorporation into sulfur-containing biomolecules. The oxidoreductase is composed of two subunits, an NADPH, FMN, and FAD-binding diflavin reductase and an iron siroheme and Fe4S4-containing oxidase. How they interact has been an unknown for over 50 years because the complex is highly flexible, thus has been intransigent for traditional X-ray or cryo-EM structural analysis. Using a combination of the chameleon plunging system with a fluorinated lipid we overcame the challenge of preserving the minimal dimer between the subunits for high-resolution cryo-EM analysis. Here, we report the first structure of the complex between the reductase and oxidase, revealing how they interact in a minimal interface. Further, we determined the structural elements that discriminate between the pairing of a siroheme-containing oxidase with a diflavin reductase or a ferredoxin partner to channel the six electrons that reduce sulfite to sulfide.

[Uric Acid Metabolism, Uric Acid Transporters and Dysuricemia].

Serum urate levels are determined by the balance between uric acid production and uric acid excretion capacity from the kidneys and intestinal tract. Dysuricemia, including hyperuricemia and hypouricemia, develops when the balance shifts towards an increase or a decrease in the uric acid pool. Hyperuricemia is mostly a multifactorial genetic disorder involving several disease susceptibility genes and environmental factors. Hypouricemia, on the other hand, is caused by genetic abnormalities. The main genes involved in dysuricemia are xanthine oxidoreductase, an enzyme that produces uric acid, and the urate transporters urate transporter 1/solute carrier family 22 member 12 (URAT1/SLC22A12), glucose transporter 9/solute carrier family 2 member 9 (GLUT9/SLC2A9) and ATP binding cassette subfamily G member 2 (ABCG2). Deficiency of xanthine oxidoreductase results in xanthinuria, a rare disease with marked hypouricemia. Xanthinuria can be due to a single deficiency of xanthine oxidoreductase or in combination with aldehyde oxidase deficiency as well. The latter is caused by a deficiency in molybdenum cofactor sulfurase, which is responsible for adding sulphur atoms to the molybdenum cofactor required for xanthine oxidoreductase and aldehyde oxidase to exert their action. URAT1/SLC22A12 and GLUT9/SLC2A9 are involved in urate reabsorption and their deficiency leads to renal hypouricemia, a condition that is common in Japanese due to URAT1/SLC22A12 deficiency. On the other hand, ABCG2 is involved in the secretion of urate, and many Japanese have single nucleotide polymorphisms that result in its reduced function, leading to hyperuricemia. In particular, severe dysfunction of ABCG2 leads to hyperuricemia with reduced extrarenal excretion.