Role of histone H3K4 methyltransferase in regulating Monascus pigments production by red light-coupled magnetic field.

Light, magnetic field, and methylation affected the growth and secondary metabolism of fungi. The regulation effect of the three factors on the growth and Monascus pigments (MPs) synthesis of Monascus purpureus was investigated in this study. 5-azacytidine (5-AzaC), DNA methylation inhibitor, was used to treat M. purpureus (wild-type, WT). Twenty micromolar 5-AzaC significantly promoted the growth, development, and MPs yield. Moreover, 250 lux red light and red light coupled magnetic field (RLCMF) significantly promoted the biomass. For WT, red light, and RLCMF significantly promoted MPs yield. But compared with red light treatment, only 0.2 mT RLCMF promoted the alcohol-soluble MPs yield. For histone H3K4 methyltransferase complex subunit Ash2 gene knockout strain (ΔAsh2), only 0.2 mT RLCMF significantly promoted water-soluble MPs yield. Yet red light, 1.0 and 0.2 mT RLCMF significantly promoted alcohol-soluble MPs yield. This indicated that methylation affected the MPs biosynthesis. Red light and weaker MF had a synergistic effect on the growth and MPs synthesis of ΔAsh2. This result was further confirmed by the expression of related genes. Therefore, histone H3K4 methyltransferase was involved in the regulation of the growth, development, and MPs synthesis of M. purpureus by the RLCMF.

Life-History and Ecological Correlates of Egg and Clutch Mass Variation in Sympatric Bird Species at High Altitude.

The variation in egg and clutch mass in sympatric species at high altitudes is poorly understood, and the potential causes of variation are rarely investigated. This study aimed to describe the interspecific variation in avian egg and clutch mass among 22 sympatric bird species at an altitude of 3430 m. Our objective was to reduce potential confounding effects of biotic/abiotic factors and investigated hypotheses concerning allometry, clutch size, parental care, nest predation, and lifespan as possible correlates and explanations for the observed variation. Our findings indicated that both egg and clutch mass evolve with body mass across species. We found that egg mass variation was not explained by clutch size when controlling for allometric effects, which contrasts the "egg mass vs. clutch size trade-off" hypothesis. Additionally, we found that clutch mass was positively associated with parental care (reflected by development period) but negatively associated with predation rate. By substituting egg mass and clutch size into the models, we found that clutch size was significantly correlated with parental care, predation rate, and lifespan, while egg mass was only significantly associated with development period. Overall, these findings support life-history theories suggesting that reduced clutch size or mass is associated with a higher risk of predation, reduced parental care, but longer adult lifespan. Interestingly, our results indicate that clutch size has a greater influence on these factors compared to egg mass. This could be attributed to the fact that smaller clutch sizes result in a more notable decrease in energetic allocation, as they require a reduced effort in terms of offspring production, incubation, and feeding, as opposed to solely reducing egg size. These findings contribute to the growing evidence that life-history and ecological traits correlate with egg and clutch mass variation in sympatric species. However, further research is needed to explore the potential evolutionary causes underlying these patterns.

Effect of enzymatic hydrolysis on volatile flavor compounds of Monascus-fermented tartary buckwheat based on headspace gas chromatography-ion mobility spectrometry.

Tartary buckwheat was hydrolyzed with α-amylase, pullulanase, α-amylase and pullulanase double enzymes and fermented by Monascus. The fermentation products were named as enzymolysis-Monascus-fermented tartary buckwheat (EMFTB). The composition and content of volatile flavor compounds in EMFTB were investigated. The results showed that α-amylase and pullulanase hydrolysis reduced starch content and raised protein, flavonoids, Monacolin K and Monascus pigments content of EMFTB. Meanwhile, double enzyme hydrolysis significantly changed the principal components of volatile substances and affected the varieties and content of volatile organic substances in EMFTB using electronic nose and headspace gas chromatography-ion mobility chromatography (HS-GC-IMS). The volatile organic substances and main aroma components increased significantly in EMFTB, including 2-heptanone, 3-methyl-1-butanol, butan-1-ol, 2-methyl-1-propanol and other substances. These results indicate that the amylase hydrolysis plays an important role in improving the flavor quality of EMFTB.

Lactic acid bacteria synergistic fermentation affects the flavor and texture of bread.

Fermentation strains play a key role in the quality of bread. The combination of yeast and lactic acid bacteria (LAB) may effectively improve the function and nutritional properties of bread. In this study, the dough was fermented to make bread by using single strain (Saccharomyces cerevisiae, mode A), the combination of two strains (S. cerevisiae and Lactiplantibacillus plantarum, mode B; S. cerevisiae and Lactobacillus delbrueckii, mode C), or three strains (S. cerevisiae, L. plantarum, and L. delbrueckii, mode D). The specific volume, texture, and aroma substances of bread were evaluated. The possibility of mixed fermentation of selected yeast and LAB to replace natural fermentation dough was evaluated. The results showed that the specific volume of bread in mode B was 15.2% higher than that of mode A. The structure was softer and the taste was more vigorous in mode B bread. The content of volatile compounds was highest in mode B bread among the four mode bread. The characteristic flavors were ethyl 2-hydroxypropionate and z-3-hexenol. The cofermentation in mode B made the bread aroma richer and gave better aroma characteristics to bread. Therefore, the fermentation of S. cerevisiae and L. plantarum can be recommended to replace naturally fermented dough to improve the quality of bread. PRACTICAL APPLICATION: L. plantarum and L. delbrueckii, separately or together, assisted in yeast fermentation to make bread. The specific volume, texture, and aroma substances of bread were evaluated to replace natural fermented dough with mixed fermentation. L. plantarum-assisted yeast fermentation improved the specific volume, texture, and aroma of bread. The characteristic flavors were ethyl 2-hydroxypropionate and z-3-hexenol in bread. Therefore, the fermentation of S. cerevisiae and L. plantarum could replace naturally fermented dough to improve the quality of bread.

A facile macroporous resin-based method for separation of yellow and orange Monascus pigments.

The yellow Monascus pigments (YMPs) named monascin and ankaflavin and the orange Monascus pigments (OMPs) named rubropunctatin and monascorubrin are two groups of bioactive components in a mixture state in the Monascus fermented products. In order to separate these two groups of bioactive pigments, a facile macroporous resin-based method was developed. The weak-polar resin CAD-40 was selected from the seven tested macroporous resins as it revealed better properties for the adsorption and desorption of the YMPs and OMPs. Then, CAD-40 resin was used for column-chromatographic separation. After eluted by 4 bed volumes of ethanol, the yellow group (monascin and ankaflavin) and the orange group (rubropunctatin and monascorubrin) were successfully separated and purified, with an increased content from 49.3% and 44.2% in the crude pigment extract to 85.2% and 83.0% in the final products, respectively. This method would be helpful for the large-scale separation and purification of Monascus pigment products with specific bioactivity.

Mutational analysis of MpPhy reveals magnetoreception and photosensitivity involvement in secondary metabolites biosynthesis in Monascus purpureus.

Light or low frequency magnetic field (LF-MF) as one of the cultivation environments affects secondary metabolites (SMs) production of M. purpureus. Phytochrome (Phy) is a hybrid histidine kinase possessing dual properties of photoreceptor and kinase to sense red and far-red light. The interaction effects of LF-MF and light on SMs of M. purpureus was investigated by knocking out the Phy-like gene in M. purpureus (MpPhy) by homologous recombination. A MpPhy-deletion (ΔMpPhy) strain produced less Monascus pigments (MPs) and monacolin K (mon K) than the wild-type (WT) strain and reduced citrinin production by 78.3% on 10th day but didn't affect the biomass. These results indicated that the MpPhy gene is involved in SMs biosynthesis of M. purpureus. MPs production in WT was decreased significantly when the inoculum was exposed to white/blue/green/red light (500 Lux). But it in ΔMpPhy was no significant difference when exposed to white/red light. The colony size of ΔMpPhy was smaller on potato dextrose agar media containing 0.01% SDS. These results indicated that the deletion of MpPhy gene affected the aerial hyphae and increased sensitivity to cell membrane stress but decreased sensitivity to red light. The inoculum of both WT and ΔMpPhy was exposure to the LF-MF (50 Hz). The accumulation of WT secondary metabolites was not changed, while SMs production of ΔMpPhy was significantly enhanced under exposed to 2.0 mT LF-MF. This indicated that the decrease of SMs caused by the deletion of MpPhy gene was restored by LF-MF. It revealed that there is a crosstalk between magnetoreception and photosensitivity.

Low-Frequency Magnetic Field of Appropriate Strengths Changed Secondary Metabolite Production and Na+ Concentration of Intracellular and Extracellular Monascus purpureus.

Monascus purpureus is used to yield edible pigments accompanied by mycotoxin-citrinin. A low-frequency (<300 Hz) magnetic field (LF-MF) affects microbial metabolism. The link of LF-MF with secondary metabolites and intracellular and extracellular Na+ levels in M. purpureus was determined. The fermentation broth was exposed to LF-MF during the first 2 days of fermentation and continuously cultured at 30°C and 200 rpm until the 8th day of fermentation. Results showed that LF-MF treatments didn't affect the growth of M. purpureus in liquid-state fermentation. Compared with the control, citrinin production showed a decrease of 45.0%, while yellow, red, and orange pigment production showed an increase of 72.9, 73.9, and 40.1%, respectively, with LF-MF treatment of 1.6 mT. This was in agreement with downregulation of pksCT and ctnA, and upregulation of pksPT, pigR, veA, and laeA at the transcriptional level. Moreover, 1.6 mT LF-MF exposure caused the transfer of Na+ from extracellular to intracellular, which was validated through the upregulation of transmembrane sensor synthesis genes and the changes in the relative expression levels of the P-type ATPase and protein phosphatase genes. This study established that LF-MF could inhibit citrinin and stimulate pigment production and change intracellular and extracellular Na+ concentrations. Bioelectromagnetics. 2020;41:289-297 © 2020 Bioelectromagnetics Society.

Lack of the Histone Methyltransferase Gene Ash2 Results in the Loss of Citrinin Production in Monascus purpureus.

ABSTRACT: Absent, small, or homeotic discs 2 (Ash2), a histone H3K4 methyltransferase complex, has been implicated in the control of hyphal development and secondary metabolism in many kinds of filamentous fungi. We constructed an Ash2 deletion mutant (ΔAsh2) by using an Agrobacterium-mediated gene knockout method to investigate the function of the Ash2 gene in the mold Monascus purpureus. Lack of the Ash2 gene resulted in the formation of a lower colony phenotype with fluffy aerial hyphae that autolyzed as the colony grew on potato dextrose agar at 30°C. The production of pigments and the number of conidia were significantly lower in the ΔAsh2 than in the wild type. Citrinin production by the ΔAsh2 was not detected during 15 days of fermentation. Relative expression levels of secondary metabolite regulatory genes PigR and CTNR, secondary metabolite synthesizing genes PKSPT and CTN, key genes of mitogen-activated protein kinase pathway Spk1 and its downstream gene mam2, the conidium development control gene BrlA, and global regulatory genes LaeA and VeA were detected by the quantitative real-time PCR. These results indicate that the Ash2 gene is involved in conidial germination, pigment production, and citrinin production and plays a key role in development and secondary metabolism in M. purpureus.

Composition, diversity and function of gastrointestinal microbiota in wild red-billed choughs (Pyrrhocorax pyrrhocorax)

Hitherto, virtually nothing is known about the microbial communities related to the bird species in the family Corvidae. To fill this gap, the present study was conducted to provide a baseline description of the gut microbiota of wild red-billed choughs (Pyrrhocorax pyrrhocorax). In this study, microbiota from four gastrointestinal locations (oropharynx, gizzard, small intestine, and large intestine) of three wild red-billed choughs were analyzed using the Illumina MiSeq sequencing platform by targeting the V4-V5 regions of the 16S rRNA genes. The gut microbiota of the red-billed choughs were dominated by the phylum Firmicutes (59.56%), followed by Proteobacteria (16.56%), Bacteroidetes (13.86%), and Actinobacteria (7.03%), which were commonly detected in avian gut ecosystems. Genus-level compositions were found to be largely dominated by Lactobacillus (18.21%), Weissella (12.37%), Erysipelatoclostridium (6.94%), Bacteroides (6.63%), Escherichia-Shigella (5.15%), Leuconostoc (4.60%), Proteus (3.33%), Carnobacterium (2.71%), Lactococcus (1.69%), and Enterococcus (1.63%). The overall intestinal microbiota was enriched with functions related to ATP-binding cassette (ABC) transporters, DNA repair and recombination proteins, purine metabolism, ribosome, transcription factors, pyrimidine metabolism, peptidases, and two-component system. In terms of four different gastrointestinal locations, hierarchical clustering analysis and principal coordinate analysis showed that microbial communities of the oropharynx, gizzard, small intestine, and large intestine formed four separated clusters. A total of 825 OTUs and 382 genera were detected in all four gastrointestinal locations, which were considered as the major microbes in the intestines of red-billed choughs. Coexistence of lactic acid bacteria and potential pathogens in the gut environments of red-billed choughs required further investigations

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Acidic conditions induce the accumulation of orange Monascus pigments during liquid-state fermentation of Monascus ruber M7.

The influence of pH on the biosynthesis of orange Monascus pigments (OMPs) in Monascus ruber M7 was investigated. Under acidic fermentation conditions, pigment mixtures predominantly rich in OMPs were obtained. HPLC analysis revealed the presence of four orange components (O1-O4) and four yellow components (Y1-Y4) in the mixtures, and the dominant ones were O1 and O3, which accounted for 56.0% to 75.9% of the total pigments in the pH range 3-6. Subsequently, O1 and O3 were identified by LC-DAD-ESI/MS as Rubropunctatin and Monascorubrin, respectively. The yield of OMPs was observed to be inversely dependent on pH. At pH 3, large amounts of OMPs with high purity (79.1%) were accumulated. A real-time quantitative PCR analysis revealed that the expression of genes related to the biosynthesis of OMPs in M. ruber M7 was upregulated at acidic pH as compared to neutral pH, and the variation in the level of expression of these genes with pH was consistent with the production of OMPs. These results indicated that the large accumulation of OMPs under acidic condition involved the acidic pH-induced transcription of genes related to the biosynthesis of OMPs. These results would contribute towards the development of an efficient technology for large-scale production of OMPs.

Correction: Zhen, Z. et al. NaCl Inhibits Citrinin and Stimulates Monascus Pigments and Monacolin K Production. Toxins 2019, 11, 118.

The authors wish to make the following correction to their paper [...].

Composition, diversity and function of gastrointestinal microbiota in wild red-billed choughs (Pyrrhocorax pyrrhocorax).

Hitherto, virtually nothing is known about the microbial communities related to the bird species in the family Corvidae. To fill this gap, the present study was conducted to provide a baseline description of the gut microbiota of wild red-billed choughs (Pyrrhocorax pyrrhocorax). In this study, microbiota from four gastrointestinal locations (oropharynx, gizzard, small intestine, and large intestine) of three wild red-billed choughs were analyzed using the Illumina MiSeq sequencing platform by targeting the V4-V5 regions of the 16S rRNA genes. The gut microbiota of the red-billed choughs were dominated by the phylum Firmicutes (59.56%), followed by Proteobacteria (16.56%), Bacteroidetes (13.86%), and Actinobacteria (7.03%), which were commonly detected in avian gut ecosystems. Genus-level compositions were found to be largely dominated by Lactobacillus (18.21%), Weissella (12.37%), Erysipelatoclostridium (6.94%), Bacteroides (6.63%), Escherichia-Shigella (5.15%), Leuconostoc (4.60%), Proteus (3.33%), Carnobacterium (2.71%), Lactococcus (1.69%), and Enterococcus (1.63%). The overall intestinal microbiota was enriched with functions related to ATP-binding cassette (ABC) transporters, DNA repair and recombination proteins, purine metabolism, ribosome, transcription factors, pyrimidine metabolism, peptidases, and two-component system. In terms of four different gastrointestinal locations, hierarchical clustering analysis and principal coordinate analysis showed that microbial communities of the oropharynx, gizzard, small intestine, and large intestine formed four separated clusters. A total of 825 OTUs and 382 genera were detected in all four gastrointestinal locations, which were considered as the major microbes in the intestines of red-billed choughs. Coexistence of lactic acid bacteria and potential pathogens in the gut environments of red-billed choughs required further investigations.

NaCl Inhibits Citrinin and Stimulates Monascus Pigments and Monacolin K Production.

Applications of beneficial secondary metabolites produced by Monascus purpureus (M. purpureus) could be greatly limited for citrinin, a kidney toxin. The link of NaCl with cell growth and secondary metabolites in M. purpureus was analyzed with supplementations of different concentrations of NaCl in medium. The content of citrinin was reduced by 48.0% but the yellow, orange, red pigments and monacolin K productions were enhanced by 1.7, 1.4, 1.4 and 1.4 times, respectively, compared with those in the control using NaCl at 0.02 M at the 10th day of cultivation. NaCl didn't affect the cell growth of M. purpureus. This was verified through the transcriptional up-regulation of citrinin synthesis genes (pksCT and ctnA) and the down-regulation of the Monascus pigments (MPs) synthesis genes (pksPT and pigR). Moreover, the reactive oxygen species (ROS) levels were promoted by NaCl at the 2nd day of cultivation, and then inhibited remarkably with the extension of fermentation time. Meanwhile, the activities of superoxide dismutase (SOD) and catalase (CAT), and the contents of total glutathione (T-GSH) were significantly enhanced in the middle and late stages of cultivation. The inhibition effect on colony size and the growth of aerial mycelia was more obvious with an increased NaCl concentration. Acid and alkaline phosphatase (ACP and AKP) activities dramatically increased in NaCl treatments. NaCl could participate in secondary metabolites synthesis and cell growth in M. purpureus.

Comparative metagenomics of the gut microbiota in wild greylag geese (Anser anser) and ruddy shelducks (Tadorna ferruginea).

Gut microbiome contributes to host health by maintaining homeostasis, increasing digestive efficiency, and facilitating the development of immune system. Wild greylag geese (Anser anser) and ruddy shelducks (Tadorna ferruginea), migrating along the central Asian flyway, appear to be one of the most popular species in the rare birds rearing industries of China. However, the structure and function of the gut microbial communities associated with these two bird species remain poorly understood. Here, for the first time, we compared gut metagenomes from greylag geese to ruddy shelducks and investigated the similarities and differences between these two bird species in detail. Taxonomic classifications revealed the top three bacterial phyla, Firmicutes, Proteobacteria, and Fusobacteria, in both greylag geese and ruddy shelducks. Furthermore, between the two species, 12 bacterial genera were found to be more abundant in ruddy shelducks and 41 genera were significantly higher in greylag geese. A total of 613 genera (approximately 70%) were found to be present in both groups. Metabolic categories related to carbohydrate metabolism, metabolism of cofactors and vitamins, lipid metabolism, amino acid metabolism, and glycan biosynthesis and metabolism were significantly more abundant in ruddy shelducks, while greylag geese were enriched in nucleotide metabolism and energy metabolism. The herbivorous greylag geese gut microbiota harbored more carbohydrate-active enzymes than omnivorous ruddy shelducks. In our study, a range of antibiotic resistance categories were also identified in the gut microbiota of greylag geese and ruddy shelducks. In addition to providing a better understanding of the composition and function of wild birds gut microbiome, this comparative study provides reference values of the artificial domestication of these birds.

iTRAQ-Based Quantitative Proteomic Analysis Reveals Changes in Metabolite Biosynthesis in Monascus purpureus in Response to a Low-Frequency Magnetic Field.

BACKGROUND: Low-frequency magnetic fields (LF-MFs) dampen the citrinin output by Monascus purpureus in fermentations. The influence of LF-MFs on biosynthesis by M. purpureus was evaluated at the protein level. METHODS: Cultures were treated with a 1.6-mT MF from day 0 to day 2 of incubation, and secondary metabolite production was evaluated on the day 12 of incubation. All proteins were extracted from M. purpureus mycelia and subjected to isobaric tags for relative and absolute quantification (iTRAQ) labeling and subsequent liquid chromatography/mass spectrometry (LC-MS/MS) analysis on day 6 of fermentation. RESULTS: There was no difference in biomass between the treated samples and the control. Citrinin production was 46.7% lower, and the yields of monacolin K and yellow, orange, and red pigment were 29.3%, 31.3%, 41.7%, and 40.3% higher, respectively, in the exposed samples compared to the control. Protein expression in M. purpureus under LF-MF treatment was quantified using iTRAQ technology. Of 2031 detected proteins, 205 were differentially expressed. The differentially-expressed proteins were subjected to Gene Ontology (GO) functional annotation and statistical analysis, which revealed that they mainly refer to biological metabolism, translation, antioxidant, transport and defense pathways. Among all the tagged proteins, emphasis was placed on the analysis of those involved in the synthesis of citrinin, pigment and monacolin K was emphasized. CONCLUSIONS: LF-MFs affected Monascus secondary metabolism at the protein level, and aggregate data for all the protein profiles in LF-MF-treated Monascus was obtained.

Comparative analyses of the gut microbiota among three different wild geese species in the genus Anser.

In this study, we characterized for the first time the gut microbiota of Greylag geese (Anser anser) using high-throughput 16S rRNA gene sequencing technology. The results showed that the phyla Firmicutes (78.55%), Fusobacteria (9.38%), Proteobacteria (7.55%), Bacteroidetes (1.82%), Cyanobacteria (1.44%), and Actinobacteria (0.61%) dominated the gut microbial communities in the Greylag geese. Then, the variations of gut microbial community structures and functions among the three geese species, Greylag geese, Bar-headed geese (Anser indicus), and Swan geese (Anser cygnoides), were explored. The greatest gut microbial diversity was found in Bar-headed geese group, while other two groups had the least. The dominant bacterial phyla across all samples were Firmicutes and Proteobacteria, but several characteristic bacterial phyla and genera associated with each group were also detected. At all taxonomic levels, the microbial community structure of Swan geese was different from those of Greylag geese and Bar-headed geese, whereas the latter two groups were less different. Functional KEGG categories and pathways associated with carbohydrate metabolism, energy metabolism, and amino acid metabolism were differentially expressed among different geese species. Taken together, this study could provide valuable information to the vast, and yet little explored, research field of wild birds gut microbiome.

Mycothiol protects Corynebacterium glutamicum against acid stress via maintaining intracellular pH homeostasis, scavenging ROS, and S-mycothiolating MetE.

Mycothiol (MSH) plays a major role in protecting cells against oxidative stress and detoxification from a broad range of exogenous toxic agents. In the present study, we reveal that intracellular MSH contributes significantly to the adaptation to acidic conditions in the model organism Corynebacterium glutamicum. We present evidence that MSH confers C. glutamicum with the ability to adapt to acidic conditions by maintaining pHi homeostasis, scavenging reactive oxygen species (ROS), and protecting methionine synthesis by the S-mycothiolation modification of methionine synthase (MetE). The role of MSH in acid adaptation was further confirmed by improving the acid tolerance of C. glutamicum by overexpressing the key MSH synthesis gene mshA. Hence, our work provides insights into a previously unknown, but important, aspect of the C. glutamicum cellular response to acid stress. The results reported here may help to understand acid tolerance mechanisms in acid sensitive bacteria and may open a new avenue for improving acid resistance in industry strains for the production of bio-based chemicals from renewable biomass.

Mechanism for the primary transformation of acetaminophen in a soil/water system.

The transformation of acetaminophen (APAP) in a soil/water system was systematically investigated by a combination of kinetic studies and a quantitative analysis of the reaction intermediates. Biotransformation was the predominant pathway for the elimination of APAP, whereas hydrolysis or other chemical transformation, and adsorption processes made almost no contribution to the transformation under a dark incubation. Bacillus aryabhattai strain 1-Sj-5-2-5-M, Klebsiella pneumoniae strain S001, and Bacillus subtilis strain HJ5 were the main bacteria identified in the biotransformation of APAP. The soil-to-water ratio and soil preincubation were able to alter the transformation kinetic pattern. Light irradiation promoted the overall transformation kinetics through enhanced biotransformation and extra photosensitized chemical reactions. The transformation pathways were strongly dependent on the initial concentration of APAP. The main primary transformation products were APAP oligomers and p-aminophenol, with the initial addition of 26.5 and 530 µM APAP, respectively. APAP oligomers accounted for more than 95% of transformed APAP, indicating that almost no bound residues were generated through the transformation of APAP in the soil/water system. The potential environmental risks of APAP could increase following the transformation of APAP in the soil/water system because of the higher toxicity of the transformation intermediates.

Functional characterization of a mycothiol peroxidase in Corynebacterium glutamicum that uses both mycoredoxin and thioredoxin reducing systems in the response to oxidative stress.

Previous studies have identified a putative mycothiol peroxidase (MPx) in Corynebacterium glutamicum that shared high sequence similarity to sulfur-containing Gpx (glutathione peroxidase; CysGPx). In the present study, we investigated the MPx function by examining its potential peroxidase activity using different proton donors. The MPx degrades hydrogen peroxide and alkyl hydroperoxides in the presence of either the thioredoxin/Trx reductase (Trx/TrxR) or the mycoredoxin 1/mycothione reductase/mycothiol (Mrx1/Mtr/MSH) regeneration system. Mrx1 and Trx employ different mechanisms in reducing MPx. For the Mrx1 system, the catalytic cycle of MPx involves mycothiolation/demycothiolation on the Cys(36) sulfenic acid via the monothiol reaction mechanism. For the Trx system, the catalytic cycle of MPx involves formation of an intramolecular disulfide bond between Cys(36) and Cys(79) that is pivotal to the interaction with Trx. Both the Mrx1 pathway and the Trx pathway are operative in reducing MPx under stress conditions. Expression of mpx markedly enhanced the resistance to various peroxides and decreased protein carbonylation and intracellular reactive oxygen species (ROS) accumulation. The expression of mpx was directly activated by the stress-responsive extracytoplasmic function-σ (ECF-σ) factor [SigH]. Based on these findings, we propose that the C. glutamicum MPx represents a new type of GPx that uses both mycoredoxin and Trx systems for oxidative stress response.

Enhancing Corynebacterium glutamicum robustness by over-expressing a gene, mshA, for mycothiol glycosyltransferase.

Over-expression of the gene, mshA, coding for mycothiol glycosyl transferase improved the robustness of Corynebacterium glutamicum to various stresses. Intracellular mycothiol (MSH) content was increased by 114 % in WT(pXMJ19-mshA) compared to WT(pXMJ19). Survival rates increased by 44, 39, 90, 77, 131, 87, 52, 47, 57, 85 and 33 % as compared to WT(pXMJ19) under stress by H2O2 (40 mM), methylglyoxal (5.8 mM), erythromycin (0.08 mg ml(-1)), streptomycin (0.005 mg ml(-1)), Cd(2+) (0.01 mM), Mn(2+) (2 mM), formic acid (0.05 %), acetic acid (0.15 %), levulinic acid (0.25 %), furfural (7.2 mM), and ethanol (10 % v/v), respectively. Increased MSH content also decreased the concentration of reactive oxygen species in the presence of the above stresses. Our results may open a new avenue for enhancing robustness of industrial bacteria for production of commodity chemicals.