Corynebacterium glutamicum is an important industrial strain for amino acids and a key model organism for human pathogens. The study of C. glutamicum oxidoreductases, such as mycoredoxin 1 (Mrx1), dithiol-disulfide isomerase DsbA, and DsbA-like Mrx1, is helpful for understanding the survival, pathogenic infection, and stress resistance of its homologous species. However, the action mode and enzymatic function of C. glutamicum NCgl0018 preserving the Cys-Pro-Phe-Cys motif, annotated as a putative DsbA, have remained enigmatic. Here, we report that the NCgl0018-deleted strain increased sensitivity to various oxidative stresses. The ncgl0018 expression was induced in the stress-responsive extracytoplasmic function-sigma (ECF-σ) factor SigH- and organic peroxide- and antibiotic-sensing regulator (OasR)-dependent manner by stress. NCgl0018 reduced S-mycothiolated mixed disulfides and intramolecular disulfides via a monothiol-disulfide mechanism preferentially linking the mycothiol/mycothione reductase/NADPH electron pathway. Site-directed mutagenesis confirmed Cys107 was the resolving Cys residue, while Cys104 was the nucleophilic cysteine that was oxidized to a sulfenic acid and then could form an intramolecular disulfide bond with Cys107 or a mixed disulfide with mycothiol under stress. Biochemical analyses indicated that NCgl0018 lacked oxidase properties like the classical DsbA. Further, enzymatic rates and substrate preferences of NCgl0018 were highly similar to those of DsbA-like Mrx1. Collectively, our study presented the first evidence that NCgl0018 protected against stresses by functioning as a novel DsbA-like Mrx1 but not DsbA and Mrx1.
Streptomycetes are characterized by their ability to produce structurally diverse compounds as secondary metabolites and by their complex developmental life cycle, which includes aerial mycelium formation and sporulation. The production of secondary metabolites is growth-stage dependent, and generally coincides with morphological development on a solid culture. Streptomyces sp. BB47 produces several types of bioactive compounds and displays a bald phenotype that is devoid of an aerial mycelium and spores. Here, we demonstrated by genome analysis and gene complementation experiments that the bald phenotype arises from the bldA gene, which is predicted to encode the Leu-tRNAUUA molecule. Unlike the wild-type strain producing jomthonic acid A (1) and antarlide A (2), the strain complemented with a functional bldA gene newly produced milbemycin (3). The chemical structure of compound 3 was elucidated on the basis of various spectroscopic analyses, and was identified as milbemycin A4, which is an insecticidal/acaricidal antibiotic. These results indicate that genetic manipulation of genes involved in morphological development in streptomycetes is a valuable way to activate cryptic biosynthetic pathways.
In this study, two fibrous carbon anodes (namely, pleated non-woven graphite (PNWG) and carbon brush (CB) made from artificial carbon) and bamboo charcoal (BC) were evaluated for current recovery from sewage wastewater. When these anodes were polarized at 0.2 V vs. Ag/AgCl in sewage wastewater, CB produced a maximum current of 2.9 A/m2. This exceeded that produced by PNWG (1.5 A/m2) and BC (1.4 A/m2). The accumulative charge recovery achieved with CB was superior to those achieved with the other two (1.6- and 2.2-fold higher than that with PNWG and BC, respectively). During the cyclic voltammetry analysis, CB demonstrated the highest catalytic current with maximum potential in the range of –0.6 to 0.4 V vs. Ag/AgCl and the smallest anode resistance (0.20 Ωm2). Direct cell counting revealed that the fibrous anodes (CB and PNWG) attached most of the cells in the anodes (80%), whereas BC did not. In contrast, the proportion of Geobacter species, a representative electrogenic microorganism in the total bacteria, was observed to be similar among the three anodes (4.4–5.8%). The tubular microbial fuel cell (ø 5.0 cm) equipped with an air-chamber core wrapped with an anion exchange membrane (AEM) and the CB delivered a current of 1.8 A/m2. This is higher than those reported in the existing literature for the same microbial fuel cell (MFC) configuration. This indicates that the alteration of the anode from planar to brush can contribute toward improving the current recovery through the air-cathode-AEM-MFC. The BC needs improvement to have more specific surface area, whereas it showed superiority in cost efficiency considering material and processing.
During the making of rice-koji for sake production, 2,4,6-trichlorophenol (TCP) is O-methylated to 2,4,6-trichloroanisole (TCA) by the koji-mold, Aspergillus oryzae, resulting in a musty/moldy off-odor, which significantly reduces the quality of sake. Thus, we aim to develop A. oryzae strains with a less-efficient ability to produce TCA. TCP is a fungicide that suppresses the growth of fungi, whereas TCA does not. The exact effects of TCP on the growth of A. oryzae are unknown. However, it is assumed that a strain with low TCP conversion ability will be sensitive to TCP concentration. In this study, we investigated the effects of the different concentrations of TCP on the growth suppression of A. oryzae. As the TCP concentration in the media increased, the growth rate, and conidia formation of A. oryzae slowed down. No growth was observed in liquid culture (for 1 day at 30°C) containing more than 30 μg/mL of TCP and in agar culture (for 7 days at 30°C) containing more than 50 μg/mL of TCP. However, A. oryzae was able to grow on alpha rice containing higher concentrations of TCP. The results in agar culture are consistent with the effects of TCP on other Aspergillus species.
The terrestrial cyanobacterium Nostoc commune has a cosmopolitan distribution. It is edible, and dry thalli are sold as a food in China under the name of Di Pi Cai. The pigment composition and the genotypes were characterized to identify the cyanobacterium Di Pi Cai from China as N. commune. Myxol glycosides and ketocarotenoids were detected, as expected in Nostoc sp., but β-carotene and hydroxylated carotenoids were not detected. Nostoc-756, mycosporine-2-(4-deoxygadusoyl-ornitine), was found to be a main mycosporine-like amino acid, which indicates that Di Pi Cai belongs to the N. commune chemotype C. However, the 16S rRNA gene and the petH gene encoding ferredoxin-NADP+ oxidoreductase of Di Pi Cai did not exactly match those of genotype C found in Japan. These results suggest the unique molecular genetic features of Di Pi Cai and the global diversity of N. commune.
We describe a novel expression cassette that enables efficient and constitutive expression of the ZZ domain derived from Staphylococcus aureus protein A on the yeast cell surface to easily prepare yeast-based immunosorbents. Using this expression cassette containing the PGK1 promoter, a secretion signal derived from α-factor, and a Flo1-derived anchor protein, we successfully created a yeast-based immunosorbent for human serum albumin.
Glabridin (Glb), a polyphenolic flavonoid inhibits the growth of MDRSA (Multidrug resistant S. aureus) 4627 by inducing ROS. Glb in combination with Norfloxacin (Nor) synergistically induced oxidative stress. Increased ROS/RNS levels, in particular, affected macromolecules' (DNA, lipid, protein) integrity and distorted cell morphology. We found correlation between drug-effects and up-/down-regulation of oxidative stress-related as well as MDR genes. These findings could considerably potentiate the dosing routine of Nor in combination with Glb, which holds a promising prospective as a antibacterial agent against S. aureus.