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Xanthine oxidoreductase is a metalloenzyme that catalyzes the final steps in purine metabolism by converting hypoxanthine to xanthine and then uric acid. Allopurinol, an analog of hypoxanthine, is widely used as an antigout drug, as xanthine oxidoreductase-mediated metabolism of allopurinol to oxypurinol leads to oxypurinol rotation in the enzyme active site and reduction of the molybdenum Mo(VI) active center to Mo(IV), inhibiting subsequent urate production. However, when oxypurinol is administered directly to a mouse model of hyperuricemia, it yields a weaker urate-lowering effect than allopurinol. To better understand its mechanism of inhibition and inform patient dosing strategies, we performed kinetic and structural analyses of the inhibitory activity of oxypurinol. Our results demonstrated that oxypurinol was less effective than allopurinol both in vivo and in vitro. We show that upon reoxidation to Mo(VI), oxypurinol binding is greatly weakened, and reduction by xanthine, hypoxanthine, or allopurinol is required for reformation of the inhibitor-enzyme complex. In addition, we show oxypurinol only weakly inhibits the conversion of hypoxanthine to xanthine and is therefore unlikely to affect the feedback inhibition of de novo purine synthesis. Furthermore, we observed weak allosteric inhibition of purine nucleoside phosphorylase by oxypurinol which has potentially adverse effects for patients. Considering these results, we propose the single-dose method currently used to treat hyperuricemia can result in unnecessarily high levels of allopurinol. While the short half-life of allopurinol in blood suggests that oxypurinol is responsible for enzyme inhibition, we anticipate multiple, smaller doses of allopurinol would reduce the total allopurinol patient load.

The conversion of xanthine dehydrogenase (XDH) to xanthine oxidase (XO) occurs only in mammalian species. In fresh bovine milk, the enzyme exists predominantly as the oxidase form, in contrast to various normal organs where it is found primarily as the dehydrogenase: the mechanism of conversion to the oxidase in milk remains obscure. A systematic search for the essential factors for conversion from XDH to XO has been performed within fresh bovine milk using the highly purified dehydrogenase form after removal endogenous oxidase form by fractionation analysis. We find that conversion to the oxidase form requires four components under air: lactoperoxidase (LPO), XDH, SCN-, and substrate hypoxanthine or xanthine; the contribution of sulfhydryl oxidase appears to be minor. Disulfide bond formation between Cys-535 and Cys-995 is principally involved in the conversion, consistent with the result obtained from previous work with transgenic mice. In vitro reconstitution of LPO and SCN- results in synergistic conversion of the dehydrogenase to the oxidase the presence of xanthine, indicating the conversion is autocatalytic. Milk from an LPO knockout mouse contains a significantly greater proportion of the dehydrogenase form of the enzyme, although some oxidase form is also present, indicating that LPO contributes principally to the conversion, but that sulfhydryl oxidase (SO) may also be involved to a minor extent. All the components XDH/LPO/SCN- are necessary to inhibit bacterial growth in the presence of xanthine through disulfide bond formation in bacterial protein(s) required for replication, as part of an innate immunity system in mammals. Human GTEx Data suggest that mRNA of XDH and LPO are highly co-expressed in the salivary gland, mammary gland, mucosa of the airway and lung alveoli, and we have confirmed these human GTEx Data experimentally in mice. We discuss the possible roles of these components in the propagation of SARS-CoV-2 in these human cell types.

Sudden unexpected death in epilepsy (SUDEP) refers to unpredictable demise of a person following a seizure. Electroencephalograms can directly measure electrical activity in the brain; however, it cannot predict when seizures will occur. The use of electrocardiograms (ECGs) to monitor changes in brain electrical activity has gained attention, recently. In this case report, we retrospectively reviewed ECGs taken before and after seizure activity in a 75‐year‐old male who had a remote subarachnoid hemorrhage. Interestingly, U‐waves appeared prior to his seizures and disappeared afterward, which suggests ECGs can be used to predict epilepsy in a certain population. [ABSTRACT FROM AUTHOR]

This paper describes a method for generating toolpaths based on machining strategies for five-axis controlled machining using special tools. Traditionally, most toolpath generation studies focused on ball-end mills, proposing strategic methods to achieve high-quality machining while avoiding tool interference. Recently, special finishing tools with large cutting edge radii have gained interest for achieving higher machining efficiency. These special tools can produce smooth finished surfaces even with large pick-feed widths, leading to higher productivity. However, unlike conventional machining with ball-end mills, five-axis controlled machining using special tools lacks standardized work design procedures. This study proposes a generic tool-geometry data format for defining special tool geometries and a method for generating toolpaths using this data format. This method strategically treats special tools as conventional ball-end mills. Consequently, five-axis controlled machining for new tool geometries can be achieved using existing operational procedures. To generate toolpaths, this study utilizes a two-dimensional configuration space (C-Space). For special tools with multiple cutting edge radii, the relationship between the tool posture and cutting edge contact point is clarified by mapping the cutting edge radius information onto the C-Space. By employing this mapped cutting edge information, we can determine the interference-free tool posture corresponding to the chosen cutting edge section based on the machining strategy. Finally, the paper presents machining simulations and experiments conducted to confirm the effectiveness of the proposed method. [ABSTRACT FROM AUTHOR]

Global warming has increased the growth of pathogenic Vibrio bacteria, which can cause foodborne illnesses and death. Vibrio bacteria require iron for growth and survival. They utilize a ferric ion‐binding protein (FbpA) to bind and transport Fe3+ into the cell. FbpA from Vibrio metschnikovii (Vm) is a potential target for inhibiting its growth. Rosmarinic acid (RA) can block the binding of VmFbpA to Fe3+; however, the molecular mechanism of Fe3+ binding and RA inhibition to VmFbpA is unclear. In this study, we used x‐ray crystallography to determine the Fe3+‐binding mode of VmFbpA and the mechanism of RA inhibition. The structures revealed that in the Fe3+ bound form, Fe3+ was coordinated to VmFbpA by two Tyr residues, two HCO3− ions, and two water molecules in a six‐coordinated geometry. In contrast, in the inhibitor bound form, RA was initially bound to VmFbpA following gel filtration purification, but it was hydrolyzed to danshensu (DSS), which occupied the binding site as shown in an electron density map and reverse phase chromatography (RPC) analysis. Both RA and DSS exhibited a stronger binding affinity to VmFbpA, higher Fe3+ reduction capacity, and more potent bacteriostatic effect on V. metschnikovii compared with caffeic acid (CA), another hydrolysis product of RA. These results provide insight into the mechanism of iron acquisition by V. metschnikovii and inhibition by RA and DSS. Our findings offer clues on the development of effective strategies to prevent Vibrio infections. [ABSTRACT FROM AUTHOR]