Bioscience, Biotechnology, and Biochemistry最新文献

We previously identified M.ApeKI from Aeropyum pernix K1 as a highly thermostable DNA (cytosine-5)-methyltransferase. M.ApeKI uses the type II restriction-modification system (R-M system), among the best-studied R-M systems. Although endonucleases generally utilize Mg (II) as a cofactor, several reports have shown that MTases exhibit different reactions in the presence of metal ions. This study aim was to evaluate the enzymatic properties of DNA (cytosine-5)-methyltransferase M.ApeKI from archaea in the presence of metal ions. We evaluated the influence of metal ions on the catalytic activity and DNA binding of M.ApeKI. The catalytic activity was inhibited by Cu (II), Mg (II), Mn (II), and Zn (II), each at 5 m m. DNA binding was more strongly inhibited by 5 m m Cu (II) and 10 m m Zn (II). To our knowledge, this is the first report showing that DNA binding of type II MTase is inhibited by metal ions.

Strigolactones (SLs), plant-derived apocarotenoids, serve dual roles as phytohormones and rhizosphere signaling molecules. While exogenous administration of SLs to plants aids in studying their functions, the metabolic destiny of these administered SLs remains poorly elucidated. Our previous research demonstrated that among synthetic SL GR24 stereoisomers administered to cowpea (Vigna unguiculata), 2'-epi-GR24 undergoes selective reduction at the C-3',4' double bond in its D-ring. In this investigation, we isolated proteins from cowpea roots based on SL reducing activity and identified 12-oxophytodienoate reductase 3 homologs (VuOPR3s) as contributors to this reduction. Enzymatic assays conducted with recombinant proteins revealed that VuOPR3s exhibited a preference for reducing activity toward 2'S-configured SLs, including 2'-epi-GR24. This specificity for 2'S-configured SLs was congruent with that observed for orobanchol produced by cowpea and its stereoisomers. These findings suggest that exogenously administered SLs undergo enzymatic stereoselective reduction, underscoring the importance of considering stereospecificity when interpreting data obtained from SL usage.

Total syntheses of borolithochromes H1, H2, I1, and I2, the red pigments isolated from fossils of Jurassic putative red alga Solenopora jurassica, have been achieved. The naphthoquinone possessing a chiral sec-butyl side chain has been synthesized from (S)-2-methylbutanol. The Diels-Alder reaction of the chiral naphthoquinone and the previously reported diene was followed by one pot S-methylation/intramolecular Corey-Chaykovsky reaction/epoxide rearrangement to provide the benzo[gh]tetraphene skeleton. Complexation of the resulting ligand with trimethyl borate and the following O-demethylation furnished a 1:1 mixture of borolithochromes I1 and I2, which were separated by HPLC using CHIRALPAK IC® to afford optically pure borolithochromes I1 (6) and I2 (7). On the other hand, borolithochromes H1 and H2 were not separated by HPLC in our laboratory. Fortunately, the mixture of the methyl ethers of borolithochromes H1 and H2 were separated and O-demethylation with magnesium iodide furnished optically pure borolithochromes H1 (4) and H2 (5).

Protective effect of quercetin against acetaldehyde was evaluated using the cultured hepatocyte models with aldehyde dehydrogenase (ALDH) isozyme deficiency (aldh2-kd and aldh1a1-kd). The quercetin-induced cytoprotection against acetaldehyde in the ALDH1A1-deficient mutant (aldh1a1-kd) was weaker than that in the wild type. Furthermore, quercetin did not enhance the ALDH activity in aldh1a1-kd cells, suggesting that ALDH1A1 is involved in quercetin-induced cytoprotection.

Methanol reportedly stimulates citric acid (CA) production by Aspergillus niger and A. tubingensis; however, the underlying mechanisms remain unclear. Here, we elucidated the molecular functions of the citrate exporter gene cexA in relation to CA production by A. tubingensis WU-2223L. Methanol addition to the medium containing glucose as a carbon source markedly increased CA production by strain WU-2223L by 3.38-fold, resulting in a maximum yield of 65.5 g/L, with enhanced cexA expression. Conversely, the cexA-complementing strain with the constitutive expression promoter Ptef1 (strain LhC-1) produced 68.3 or 66.7 g/L of CA when cultivated without or with methanol, respectively. Additionally, strain LhC-2 harboring two copies of the cexA expression cassette produced 80.7 g/L of CA without methanol addition. Overall, we showed that cexA is a target gene for methanol in CA hyperproduction by A. tubingensis WU-2223L. Based on these findings, methanol-independent CA-hyperproducing strains, LhC-1 and LhC-2, were successfully generated.

Cerium oxide (CeO2) nanoparticles, as a metal oxide nanomaterial, are increasingly used for various industrial and biomedical applications. Although their cytotoxicity to bacteria and the associated mechanisms have attracted particular attention, the mechanisms behind their antifungal effects have remained unclear. This study investigated the antifungal properties of CeO2, focusing on Aspergillus oryzae. CeO2 inhibited fungal spore germination on solid substrates, and the effect was fungistatic rather than fungicidal. CeO2 inhibited fungal growth, especially under UV irradiation, and induced reactive oxygen species (ROS) production. Tocopherol reduced the intracellular ROS levels and the growth-inhibitory effects of CeO2, suggesting that ROS are involved in these growth-inhibitory effects. Transcriptomic analysis revealed upregulated expression of genes related to phospholipases and phosphate metabolism. CeO2 affected phosphate ion concentration in the medium, potentially influencing cellular responses. This research provided valuable insights into the antifungal effects of CeO2 application, which differ from those of conventional photocatalysts like TiO2.

L-Proline (Pro) is an essential amino acid additive in livestock and aquaculture feeds. Previously, we created a Pro overproducing Halomonas elongata HN6 by introducing an engineered salt-inducible Pro biosynthetic mCherry-proBm1AC operon and deleting a putA gene that encoded a Pro catabolic enzyme in the genome of H. elongata OUT30018. Here, we report a generation of a novel Pro overproducing H. elongata HN10 strain with improved salt tolerance and higher Pro yield by expressing the mCherry-proBm1AC operon and deleting the putA gene in the genome of a spontaneous mutant H. elongata Glutamic acid Over-Producing, which overproduces glutamic acid (Glu) that is a precursor for Pro biosynthesis. The optimal salt concentration for growth of H. elongata HN10 was found to be 7% to 8% w/v NaCl, and the average Pro yield of 166 mg/L was achieved when H. elongata HN10 was cultivated in M63 minimal medium containing 4% w/v glucose and 8% w/v NaCl.

Peroxisome proliferator-activated receptor γ (PPARγ) belongs to the nuclear receptor superfamily and is involved in the inflammatory process. Previously, we synthesized the ligands of PPARγ that possess the hybrid structure of a food-derived cinnamic acid derivative (CA) and GW9662, an irreversible PPARγ antagonist. These ligands activate the transcription of PPARγ through the covalent bond formation with the Cys285 residue of PPARγ, whereas their anti-inflammatory effect has not been examined yet. Here, we show the anti-inflammatory effect of the covalent PPARγ ligands in RAW264 cells, murine macrophage-like cells. GW9662 suppressed the production of nitric oxide (NO) stimulated by lipopolysaccharide and exerted a synergistic effect in combination with CA. The compounds bearing their hybrid structure dramatically inhibited NO production and transcription of proinflammatory cytokines. A comparison study suggested that the 2-chloro-5-nitrobenzoyl group of the ligands is important for anti-inflammation. Furthermore, we synthesized an alkyne-tagged analogue that becomes an activity-based probe for future mechanistic study.

In this paper, we describe our discovery of burnettiene A (1) as an antimalarial compound from the culture broth of Lecanicillium primulinum (current name: Flavocillium primulinum) FKI-6715 strain utilizing our original multidrug-sensitive yeast system. This polyene-decalin polyketide natural product was originally isolated as an antifungal active compound from Aspergillus burnettii. However, the antifungal activity of 1 has been revealed in only one fungal species, and the mechanism of action of 1 remains unknown. After the validation of mitochondrial function inhibitory of 1, we envisioned a new antimalarial drug discovery platform based on mitochondrial function inhibitory activity. We evaluated antimalarial activity and 1 showed antimalarial activity against Plasmodium falciparum FCR3 (chloroquine sensitive) and the K1 strain (chloroquine resistant). Our study revealed the utility of our original screening system based on a multidrug-sensitive yeast and mitochondrial function inhibitory activity for the discovery of new antimalarial drug candidates.

Cocoa extract (CE) offers several health benefits, such as antiobesity and improved glucose intolerance. However, the mechanisms remain unclear. Adipose tissue includes white adipose tissue (WAT) and brown adipose tissue. Brown adipose tissue leads to body fat reduction by metabolizing lipids to heat via uncoupling protein 1 (UCP1). The conversion of white adipocytes into brown-like adipocytes (beige adipocytes) is called browning, and it contributes to the anti-obesity effect and improved glucose tolerance. This study aimed to evaluate the effect of CE on glucose tolerance in terms of browning. We found that dietary supplementation with CE improved glucose intolerance in mice fed a high-fat diet, and it increased the expression levels of Ucp1 and browning-associated gene in inguinal WAT. Furthermore, in primary adipocytes of mice, CE induced Ucp1 expression through β3-adrenergic receptor stimulation. These results suggest that dietary CE improves glucose intolerance by inducing browning in WAT.