Metabolites from Streptomyces aureus (VTCC43181) and Their Inhibition of Mycobacterium tuberculosis ClpC1 Protein.

Tuberculosis is one of the most common infectious diseases in the world, caused by Mycobacterium tuberculosis. The outbreak of multiple drug-resistant tuberculosis has become a major challenge to prevent this disease worldwide. ClpC1 is a Clp ATPase protein of Mycobacterium tuberculosis, functioning as a chaperon when combined with the Clp complex. ClpC1 has emerged as a new target to discover anti-tuberculosis drugs. This study aimed to explore the ClpC1 inhibitors from actinomycetes, which have been known to provide abundant sources of antibiotics. Two cyclic peptides, including nocardamin (1), halolitoralin A (3), and a lactone pleurone (2), were isolated from the culture of Streptomyces aureus (VTCC43181). The structures of these compounds were determined based on the detailed analysis of their spectral data and comparison with references. This is the first time these compounds have been isolated from S. aureus. Compounds 1-3 were evaluated for their affection of ATPase activity of the recombinant ClpC1 protein. Of these compounds, halolitoralin A (1), a macrocyclic peptide, was effective for the ATPase hydrolysis of the ClpC1 protein.

Optimization of Aging Temperature and Heat-Treatment Pathways in Additively Manufactured 17-4PH Stainless Steel.

This study investigates the tensile behaviors of additively manufactured (AM) 17-4PH stainless steels heat-treated within various temperature ranges from 400 °C to 700 °C in order to identify the effective aging temperature. Despite an aging treatment of 400-460 °C increasing the retained austenite content, an enhancement of the tensile properties was achieved without a strength-ductility trade-off owing to precipitation hardening by the Cu particles. Due to the intricate evolution of the microstructure, aging treatments above 490 °C led to a loss in yield strength and ductility. A considerable rise in strength and a decrease in ductility were brought about by the increase in the fraction of precipitation-hardened martensitic matrix in aging treatments over 640 °C. The impact of heat-treatment pathways on aging effectiveness and tensile anisotropy was then examined. Direct aging at 482 °C for an hour had hardly any effect on wrought 17-4PH, but it increased the yield strength of AM counterparts from 436-457 to 588-604 MPa. A solid-solution treatment at 1038 °C for one hour resulted in a significant drop in the austenite fraction, which led to an increase in the yield (from 436-457 to 841-919 MPa) and tensile strengths (from 1106-1127 to 1254-1256 MPa) with a sacrifice in ductility. Improved strength and ductility were realized by a solid-solution followed by an aging treatment, achieving 1371-1399 MPa. The tensile behaviors of AM 17-4PH were isotropic both parallel and perpendicular to the building direction.

Ixeris dentata and Lactobacillus gasseri media protect against periodontitis through Nrf2-HO-1 signalling pathway.

Periodontitis is an infectious inflammation in the gums characterized by loss of periodontal ligaments and alveolar bone. Its persistent inflammation could result in tooth loss and other health issues. Ixeris dentata (IXD) and Lactobacillus gasseri media (LGM) demonstrated strong antioxidant activity, which may prevent oxidative and inflammatory periodontitis. Here, IXD and LGM extracts were investigated for antioxidative activity against oral discomfort and evaluated for their synergistic effect against oxidative and inflammatory periodontitis in a mouse model. IXD/LGM suppressed pro-inflammatory cytokines like interleukin (IL)-1ß, IL-6, and TNF-α. Additionally, it reduced pro-inflammatory mediators, nitric oxide, iNOS (inducible nitric oxide synthase), and COX-2 (cyclooxygenase-2) and enhanced AKT, Nrf2, and HO-1 activation. Similarly, IXD/LGM treatment elevated osteogenic proteins and mRNAs; alkaline phosphatase, collagen type 1 (COL1), osteopontin (OPN), and runt-related transcription factor 2 (RUNX2). Hematoxylin and Eosin (H&E) staining and micro-CT analysis confirm the positive impact of IXD/LGM on the periodontal structure and its associated inflammation. These findings demonstrate that IXD/LGM inhibits oxidative stress, periodontal inflammation, and its resultant alveolar bone loss in which Akt (also known as protein kinase B)-nuclear factor-erythroid 2-related factor 2 (Nrf2)-hemoxygenase-1 (HO-1) signaling is involved. Thus, IXD/LGM is a potential candidate against oxidative/inflammatory stress-associated periodontitis.

Potential oral probiotic Lactobacillus pentosus MJM60383 inhibits Streptococcus mutans biofilm formation by inhibiting sucrose decomposition.

Streptococcus mutans is known as a contributor to dental caries. In this work, Lactobacillus pentosus MJM60383 was selected for its strong antagonistic activity against S. mutans and was characterized by good oral probiotic properties including lysozyme tolerance, adhesive ability to oral cells, good aggregation (auto-aggregation, co-aggregation) ability, hydrogen peroxide production and inhibition of biofilm formation of S. mutans. L. pentosus MJM60383 also exhibited safety as a probiotic characterized by no hemolytic activity, no D-lactate production, no biogenic amine production, and susceptibility to antibiotics. Furthermore, the biofilm formation of S. mutans was also significantly inhibited by the supernatant of L. pentosus MJM60383. An anti-biofilm mechanism study revealed that sucrose decomposition and the production of water-insoluble exopolysaccharides by S. mutans were inhibited by the treatment with L. pentosus MJM60383 supernatant. Real-time PCR analysis indicated that the supernatant of L. pentosus MJM60383 significantly inhibited the mRNA expression of S. mutans glycosyltransferases, which synthesize glucan to construct biofilm architecture and mediate bacterial adherence. Our study demonstrated L. pentosus MJM60383 as a potential oral probiotic and revealed its anti-biofilm mechanism.

Identification of the inhibitory mechanism of ecumicin and rufomycin 4-7 on the proteolytic activity of Mycobacterium tuberculosis ClpC1/ClpP1/ClpP2 complex.

Ecumicin and rufomycin 4-7 disrupt protein homeostasis in Mycobacterium tuberculosis by inhibiting the proteolytic activity of the ClpC1/ClpP1/ClpP2 complex. Although these compounds target ClpC1, their effects on the ATPase activity of ClpC1 and proteolytic activity of ClpC1/ClpP1/ClpP2 vary. Herein, we explored the ClpC1 molecular dynamics with these compounds through fluorescence correlation spectroscopy. The effect of these compounds on the ATPase activity of ClpC1-cys, the recombinant protein for fluorescence labeling, and proteolytic activity of ClpC1-cys/ClpP1/ClpP2 were identical to those of native ClpC1, whereas the intermolecular dynamics of fluorescence-labelled ClpC1 were different. Treatment with up to 1 nM ecumicin increased the population of slower diffused ClpC1 components compared with ClpC1 without ecumicin. However, this population was considerably reduced when treated with 10 nM ecumicin. Rufomycin 4-7 treatment resulted in a slower diffused component of ClpC1, and the portion of this component increased in a concentration-dependent manner. Ecumicin can generate an abnormal ClpC1 component, which cannot form normal ClpC1/ClpP1/ClpP2, via two different modes. Rufomycin 4-7 only generates slower diffused ClpC1 component that is inadequate to form normal ClpC1/ClpP1/ClpP2. Overall, we demonstrate that ecumicin and rufomycin 4-7 use different action mechanisms to generate abnormal ClpC1 components that cannot couple with ClpP1/ClpP2.

Characterization of Leuconostoc mesenteroides MJM60376 as an oral probiotic and its antibiofilm activity.

Lactic acid bacteria have been widely used as probiotics for improving gut health. However, studies on oral probiotics were very limited. In this study, 67 lactic acid bacteria (LAB) were isolated from fermented food and screened for antagonistic activity against Streptococcus mutans, the causative pathogen of dental caries. Leuconostoc mesenteroides MJM60376 showed the highest antagonistic activity against S. mutans KCTC3065. L. mesenteroides MJM60376 also showed oral probiotic characteristics including weak acid production, lysozyme tolerance, adhesion to oral epithelial cell (YD-38), antibiotic susceptibility, and good coaggregation ability with S. mutans. Furthermore, the biofilm formation of S. mutans was significantly reduced when cocultured with L. mesenteroides. Scanning electron microscopy analysis showed that amounts of attached bacteria of S. mutans and network-like structures were significantly reduced by L. mesenteroides MJM60376. Cell-free supernatant (CFS) of L. mesenteroides MJM60376 also greatly inhibited biofilm formation of S. mutans from the adherent stage, the activity remained even after it was treated with catalase, trypsin, or pH neutralized. Expression levels of biofilm formation-related genes were significantly reduced in S. mutans when it was treated with the CFS of L. mesenteroides MJM60376. Therefore, L. mesenteroides MJM60376 has great potential to be used as a multifunctional ingredient.

Lactobacillus sakei MJM60958 as a Potential Probiotic Alleviated Non-Alcoholic Fatty Liver Disease in Mice Fed a High-Fat Diet by Modulating Lipid Metabolism, Inflammation, and Gut Microbiota.

Non-alcoholic fatty liver disease (NAFLD) is a common liver disease with a rapidly increasing number of cases worldwide. This study aimed to evaluate the effects of Lactobacillus sakei MJM60958 (MJM60958) on NAFLD in vitro and in vivo. In in vitro tests, MJM60958 significantly inhibited lipid accumulation by 46.79% in HepG2 cells stimulated with oleic acid and cholesterol (OA-C). Moreover, MJM60958 showed safe and probiotic characteristics in vitro. In the animal study, MJM60958 administration in a high-fat diet-induced NAFLD mouse model significantly reduced body weight and liver weight, and controlled aspartate aminotransferase (ALT), aspartate transaminase (AST), triglyceride (TG), urea nitrogen (BUN), and uric acid (UA) levels in the blood, which are features of NAFLD. Further, treatment with MJM60958 also reduced steatosis scores in liver tissues, serum leptin and interleukin, and increased serum adiponectin content. Moreover, administration of MJM60958 resulted in a significantly decreased expression of some genes and proteins which are related to lipid accumulation, such as fatty acid synthase (FAS), acetyl-CoA carboxylase (ACC), and sterol regulatory element-binding protein 1 (SREBP-1), and also upregulated genes and protein expression of lipid oxidation such as peroxisome proliferator-activated receptor alpha (PPARα) and carnitine palmitoyltransferase 1a (CPT1A). Administration of MJM60958 increased the relative abundance of specific microbial taxa such as Verrucomicrobia, which are abundant in non-NAFLD mice, and reduced Firmicutes, which are a major group in NAFLD mice. MJM60958 affected the modulation of gut microbiota and altered the strain profile of short-chain fatty acids (SCFAs) production in the cecum by reduced lactic acid and enhanced acetic acid production. Overall, MJM60958 showed potential as a probiotic that can prevent and treat NAFLD.

2'-5' oligoadenylate synthetase­like 1 (OASL1) protects against atherosclerosis by maintaining endothelial nitric oxide synthase mRNA stability.

Endothelial nitric oxide synthase (eNOS) decreases following inflammatory stimulation. As a master regulator of endothelial homeostasis, maintaining optimal eNOS levels is important during cardiovascular events. However, little is known regarding the mechanism of eNOS protection. In this study, we demonstrate a regulatory role for endothelial expression of 2'-5' oligoadenylate synthetase-like 1 (OASL1) in maintaining eNOS mRNA stability during athero-prone conditions and consider its clinical implications. A lack of endothelial Oasl1 accelerated plaque progression, which was preceded by endothelial dysfunction, elevated vascular inflammation, and decreased NO bioavailability following impaired eNOS expression. Mechanistically, knockdown of PI3K/Akt signaling-dependent OASL expression increased Erk1/2 and NF-κB activation and decreased NOS3 (gene name for eNOS) mRNA expression through upregulation of the negative regulatory, miR-584, whereas a miR-584 inhibitor rescued the effects of OASL knockdown. These results suggest that OASL1/OASL regulates endothelial biology by protecting NOS3 mRNA and targeting miR-584 represents a rational therapeutic strategy for eNOS maintenance in vascular disease.

Lactobacillus reuteri MJM60668 Prevent Progression of Non-Alcoholic Fatty Liver Disease through Anti-Adipogenesis and Anti-inflammatory Pathway.

Non-alcoholic fatty liver disease (NALFD) is a disease characterized by liver steatosis. The liver is a key organ involved in the metabolism of fat, protein, and carbohydrate, enzyme activation, and storage of glycogen, which is closely related to the intestine by the bidirectional relation of the gut-liver axis. Abnormal intestinal microbiota composition can affect energy metabolism and lipogenesis. In this experiment, we investigated the beneficial effect of Lactobacillus reuteri MJM60668 on lipid metabolism and lipogenesis. C57BL/6 mice were fed a high-fat diet (HFD) and orally administrated with MJM60668. Our results showed that mice treated with MJM60668 significantly decreased liver weight and liver/body weight ratio, without affecting food intake. Serum levels of ALT, AST, TG, TCHO, and IL-1ß in mice fed with MJM60668 were decreased compared to the HFD group. Investigation of gene and protein expression on the lipogenesis and lipid metabolism showed that the expression of ACC, FAS, and SREBP was decreased, and PPARα and CPT was increased. Furthermore, an increase of adiponectin in serum was shown in our experiment. Moreover, serum IL-1ß level was also significantly decreased in the treated mice. These results suggested that MJM60668 can strongly inhibit lipogenesis, enhance fatty acid oxidation, and suppress inflammation. Additionally, supplementation of MJM60668 increased the proportion of Akkermansiaceae and Lachnospiracea, confirming a potential improvement of gut microbiota, which is related to mucus barrier and decrease of triglycerides levels.

New Rufomycins from Streptomyces atratus MJM3502 Expand Anti-Mycobacterium tuberculosis Structure-Activity Relationships.

Four new rufomycins, compounds 1-4, named rufomycins 56, 57, 58, and 61, respectively, exhibiting new skeletal features, were obtained from Streptomyces atratus strain MJM3502 and were fully characterized. Compounds 1 and 2 possess a 4-imidazolidinone ring not previously encountered in this family of cyclopeptides, thereby resulting in a [5,17] bicyclic framework. The in vitro anti-Mycobacterium tuberculosis potency of compounds 3 and 4 is remarkable, with minimum inhibitory concentration values of 8.5 and 130 nM, respectively.

Enhanced anti-wrinkle activity of adlay bran fermented with Lactobacillus brevis MJM60390.

OBJECTIVES: The aim of this study was to screen the lactic acid bacteria for fermentation of adlay bran and evaluate the anti-wrinkle effect of fermented and non-fermented adlay bran. METHODS: Adlay bran was fermented with candidate LAB and extracted with 70% ethanol. The extracts from LAB-fermented adlay bran and non-fermented adlay bran were evaluated for the anti-wrinkle effects by measuring the hyaluronan, collagen, and elastin production in cells using ELISA kit. The molecular anti-wrinkle mechanism was investigated by RT-qPCR. Furthermore, the antioxidant activity, total phenolic and flavonoid content were also determined. RESULTS: Among the tested LAB, Lactobacillus brevis MJM60390 was selected for the highest glycosidase activity. Both extracts from adlay bran (NFAB) and L. brevis MJM60390-fermented adlay bran (LBFAB) showed anti-wrinkle effect, and LBFAB showed higher activity. Compared with control, hyaluronan production was increased by 24.73% and 59.38%, collagen production was increased by -13.08% and 34.19%, and the elastin production was increased by 29.78% and 53.73% by NFAB and LBFAB treatment, respectively. Investigation on the mRNA expression showed that LBFAB upregulated the expression of Has 2 and Has 3 and downregulate HYAL1 and HYAL2. LBFAB also upregulated the mRNA expression of COL1A1, COL1A2, ELN and inhibited the expression of collagenase and elastase. However, not all of these genes were regulated by NFAB. Furthermore, the antioxidant activity was significantly increased after fermentation, and the content of the phenolic and flavonoid compounds also increased in the LBFAB. CONCLUSIONS: In this study, we demonstrated that fermentation of adlay bran with L. brevis MJM60390 enhanced the anti-wrinkle activity through increasing the hyaluronan synthesis in keratinocytes and improving collagen and elastin production in dermal fibroblasts.

Discovery of Novel Iminosugar Compounds Produced by Lactobacillus paragasseri MJM60645 and Their Anti-Biofilm Activity against Streptococcus mutans.

The oral cavity contains a number of microbes. They interact with each other and play an important role in human health. Among oral cariogenic microbes, Streptococcus mutans is recognized a major etiological bacteria of dental caries. Lactobacilli strains have been promoted as possible probiotic agents against S. mutans. However, their inhibitory mechanism has not been well elucidated yet. In the present study, two new compounds with strong antibiofilm activities were purified from the culture supernatant of Lactobacillus paragasseri MJM60645, which was isolated from the human oral cavity. These compounds showed strong inhibitory activities against S. mutans biofilm formation, with IC50 (concentration at which 50% biofilm was inhibited) of 30.4 µM for compound 1 and 18.9 µM for compound 2. However, these compounds did not show bactericidal activities against S. mutans. Structure elucidation by nuclear magnetic resonance (NMR) and mass spectrometry showed that compound 1 was composed of two arabinofuranose iminosugars jointed with one glycerol and oleic acid, and compound 2 was composed of two arabinofuranose iminosugars jointed with one glycerol and nervonic acid. To the best of our knowledge, these structures were discovered for the first time in this study. Treatment of S. mutans with compound 1 strongly downregulated expression levels of genes related to biofilm formation, including gtfB, gtfC, gtfD, gbpB, brpA, spaP, ftf, and smu0630 without affecting the expression of comDE or relA. This study provides new insights into novel molecules produced by Lactobacillus to regulate the pathogenesis of S. mutans, facilitating a better understanding of the mechanism for interactions between Lactobacillus and S. mutans. IMPORTANCE In this study, we isolated lactic acid bacteria that inhibit streptococcal biofilm from the oral cavity of infants and identified two novel compounds from the supernatant of their culture broth. The two compounds are structurally similar, and both consist of iminosugars, glycerol, and unsaturated fatty acid. A search of the SciFinder database revealed that these structures are novel and were discovered for the first time in this study. Mechanism studies have shown that these compounds can inhibit the expression of biofilm synthesis-related genes. This is the first report that lactic acid bacteria inhibit streptococcal biofilms by small molecules with new chemical structures. This study not only expands the understanding of natural products derived from lactic acid bacteria but also provides a new paradigm for the understanding of the interaction of bacteria in the oral microbiota.

The antioxidant enzyme Peroxiredoxin-1 controls stroke-associated microglia against acute ischemic stroke.

Ischemic stroke is the leading cause of immortal disability and death worldwide. For treatment in the acute phase, it is necessary to control excessive reactive oxygen species (ROS) damage during ischemia/reperfusion (I/R). Microglia are well known to be closely associated with excessive ROS response in the early stage of I/R. However, the precise roles of microglia associated with mitigating ROS damage, and molecular markers of heterogenetic microglia in the I/R damaged brain has not been clarified. Here, we identified a new type of microglia associated with stroke in the I/R injured brain. Single-cell RNA sequencing (scRNA-seq) was used to assess transcriptional changes of microglia and immune cells in the contralateral (CL) and ipsilateral (IL) hemispheres after transient middle cerebral artery occlusion (tMCAO) surgery to mimic ischemic stroke. We classified a unique type of microglia with enhanced antioxidant function and markers similar to those of disease-associated microglia (DAM), designated them as stroke-associated microglia (SAM). The representative antioxidant enzyme, Peroxiredoxin-1 (Prdx1), was predominantly expressed in SAM and mediated ROS defense genes, including Txn1, Srx1, Mt1, and Mt2. In the Prdx1-/- I/R damaged brain, we observed significantly increased infarction, as assessed by TTC staining, and FACS analysis detected severe microglial cell death. Importantly, scRNA transcriptomics data showed that the SAM population was specifically decreased in Prdx1-/- mice and that these mice exhibited decreased ROS damage resistance. Inflammatory responses which were detected by ELISA and qPCR, were also increased in Prdx1-/- IL hemispheres. Finally, Prdx1-dependent antioxidative SAM were found to be essential for increasing the transcription levels of stroke-protective molecules, such as osteopontin and ferritin. A novel microglia type (SAM) is specifically activated in response to stroke I/R injury, and that Prdx1 expression is required for the activation and enhanced antioxidant function of SAM.

Potential Probiotic Lacticaseibacillus paracasei MJM60396 Prevents Hyperuricemia in a Multiple Way by Absorbing Purine, Suppressing Xanthine Oxidase and Regulating Urate Excretion in Mice.

Hyperuricemia is a metabolic disorder caused by increased uric acid (UA) synthesis or decreased UA excretion. Changes in eating habits have led to an increase in the consumption of purine-rich foods, which is closely related to hyperuricemia. Therefore, decreased purine absorption, increased UA excretion, and decreased UA synthesis are the main strategies to ameliorate hyperuricemia. This study aimed to screen the lactic acid bacteria (LAB) with purine degrading ability and examine the serum UA-lowering effect in a hyperuricemia mouse model. As a result, Lacticaseibacillus paracasei MJM60396 was selected from 22 LAB isolated from fermented foods for 100% assimilation of inosine and guanosine. MJM60396 showed probiotic characteristics and safety properties. In the animal study, the serum uric acid was significantly reduced to a normal level after oral administration of MJM60396 for 3 weeks. The amount of xanthine oxidase, which catalyzes the formation of uric acid, decreased by 81%, and the transporters for excretion of urate were upregulated. Histopathological analysis showed that the damaged glomerulus, Bowman's capsule, and tubules of the kidney caused by hyperuricemia was relieved. In addition, the impaired intestinal barrier was recovered and the expression of tight junction proteins, ZO-1 and occludin, was increased. Analysis of the microbiome showed that the relative abundance of Muribaculaceae and Lachnospiraceae bacteria, which were related to the intestinal barrier integrity, was increased in the MJM60396 group. Therefore, these results demonstrated that L. paracasei MJM60396 can prevent hyperuricemia in multiple ways by absorbing purines, decreasing UA synthesis by suppressing xanthine oxidase, and increasing UA excretion by regulating urate transporters.

Molecular epidemiological study of the G protein of human respiratory syncytial virus detected in patients with acute respiratory infections in Gyeonggi Province, South Korea.

To investigate the molecular characteristics of human respiratory syncytial virus (HRSV) detected in Gyeonggi Province from 2015/16 to 2017/18, 2331 specimens from patients with sporadic acute respiratory illness and 85 specimens from four HRSV outbreaks in the postpartum care center were analyzed by real-time reverse transcription PCR. HRSVs were detected in 97 of the 2416 (4.0%) specimens, and among the positive specimens, 38 (39.2%) were identified as HRSV-A and 59 (60.8%) as HRSV-B. During the study periods, HRSV-B predominated in all seasons, except in 2016/17 during which HRSV-A predominated. Depending on the age groups, HRSV prevalence was the highest in 0- to 2-year-old patients. Comparison of noninfected subjects with HRSV-infected subjects revealed that HRSV infection more frequently resulted in fever, nasal obstruction, and wheezing, although the frequency of sore throat was low; however, comparison of the symptoms between HRSV-A- and HRSV-B-infected patients revealed no significant differences in symptoms. Phylogenetic analysis showed that all HRSV-A patients had an ON1 genotype, and all HRSV-B patients had an BA9 genotype. These results provide a valuable reference regarding the circulating pattern and molecular characterization of HRSV. Continuous monitoring will be essential to detect newly emerging HRSV genotypes.

Lactobacillus pentosus MJM60383 Inhibits Lipid Accumulation in Caenorhabditis elegans Induced by Enterobacter cloacae and Glucose.

Gut microbiota are known to play an important role in obesity. Enterobacter cloacae, a Gram-negative bacterium, has been considered a pathogenic bacterium related to obesity in the gut. In this study, we established an obesity model of C. elegans by feeding E. cloacae combined with a high glucose diet (HGD), which significantly induced lipid accumulation. An anti-lipid mechanism study revealed that the fatty acid composition and the expression level of fat metabolism-related genes were altered by feeding E. cloacae to C. elegans under HGD conditions. Lactic acid bacteria that showed antagonistic activity against E. cloacae were used to screen anti-obesity candidates in this model. Among them, L. pentosus MJM60383 (MJM60383) showed good antagonistic activity. C. eleans fed with MJM60383 significantly reduced lipid accumulation and triglyceride content. The ratio of C18:1Δ9/C18:0 was also changed in C. elegans by feeding MJM60383. In addition, the expression level of genes related to fatty acid synthesis was significantly decreased and the genes related to fatty acid ß-oxidation were up-regulated by feeding MJM60383. Moreover, MJM60383 also exhibited a high adhesive ability to Caco-2 cells and colonized the gut of C. elegans. Thus, L. pentosus MJM60383 can be a promising candidate for anti-obesity probiotics. To the best of our knowledge, this is the first report that uses E. cloacae combined with a high-glucose diet to study the interactions between individual pathogens and probiotics in C. elegans.

Rufomycins or Ilamycins: Naming Clarifications and Definitive Structural Assignments.

Rufomycin and ilamycin are synonymous for the same class of cyclopeptides, currently encompassing 33 structurally characterized isolates and 9 semisynthetic derivatives. Elucidation of new structures prioritized the consolidation of the names and established the structures of four diastereoisomeric rufomycins with a 2-piperidinone, named rufomycins 4-7, including full 1H/13C NMR assignments. The characteristic HSQC cross-peak for the CH-5, the hemiaminal carbon in amino acid #5, allows assignment of the stereocenters C-4 and C-5 within this ring. Semisynthetic derivatives (rufomycinSS 1, 2, and 3) were prepared from a rufomycins 4 and 6 mixture to validate the structural assignments. Based on the X-ray crystal structures of rufomycins 2 and 4, considering the NMR differences of rufomycins 7 vs 4-6 compared to rufomycinSS 1 vs 2 and 3, and taking into account that two major conformers, A and B, occur in both rufomycinSS 2 and 3, structural modeling was pursued. Collectively, this paper discusses the NMR spectroscopic differences of the stereoisomers and their possible 3D conformers and correlates these with the anti-Mycobacterium tuberculosis activity. In addition, a look at the history prioritizes names and numbering schemes for this group of antibiotics and leads to consolidated nomenclature for all currently known members, natural and semisynthetic derivatives, and serves to accommodate future discoveries.

SCO6992, a Protein with ß-Glucuronidase Activity, Complements a Mutation at the absR Locus and Promotes Antibiotic Biosynthesis in Streptomyces coelicolor.

Streptomyces coelicolor is a filamentous soil bacterium producing several kinds of antibiotics. S. coelicolor abs8752 is an abs (antibiotic synthesis deficient)-type mutation at the absR locus; it is characterized by an incapacity to produce any of the four antibiotics synthesized by its parental strain J1501. A chromosomal DNA fragment from S. coelicolor J1501, capable of complementing the abs- phenotype of the abs8752 mutant, was cloned and analyzed. DNA sequencing revealed that two complete ORFs (SCO6992 and SCO6993) were present in opposite directions in the clone. Introduction of SCO6992 in the mutant strain resulted in a remarkable increase in the production of two pigmented antibiotics, actinorhodin and undecylprodigiosin, in S. coelicolor J1501 and abs8752. However, introduction of SCO6993 did not show any significant difference compared to the control, suggesting that SCO6992 is primarily involved in stimulating the biosynthesis of antibiotics in S. coelicolor. In silico analysis of SCO6992 (359 aa, 39.5 kDa) revealed that sequences homologous to SCO6992 were all annotated as hypothetical proteins. Although a metalloprotease domain with a conserved metal-binding motif was found in SCO6992, the recombinant rSCO6992 did not show any protease activity. Instead, it showed very strong ß-glucuronidase activity in an API ZYM assay and toward two artificial substrates, p-nitrophenyl-ß-D-glucuronide and AS-BI-ß-D-glucuronide. The binding between rSCO6992 and Zn2+ was confirmed by circular dichroism spectroscopy. We report for the first time that SCO6992 is a novel protein with ß-glucuronidase activity, that has a distinct primary structure and physiological role from those of previously reported ß-glucuronidases.

Antithrombotic Effect of the Ethanol Extract of Angelica gigas Nakai (AGE 232).

Cardiovascular diseases, such as stroke, are the most common causes of death in developed countries. Ischemic stroke accounts for 85% of the total cases and is caused by abnormal thrombus formation in the vessels, causing deficient blood and oxygen supply to the brain. Prophylactic treatments include the prevention of thrombus formation, of which the most used is acetylsalicylic acid (ASA); however, it is associated with a high incidence of side effects. Angelica gigas Nakai (AG) is a natural herb used to improve blood circulation via anti-platelet aggregation, one of the key processes involved in thrombus formation. We examined the antithrombotic effects of AGE 232, the ethanol extract of A. gigas Nakai. AGE 232 showed a significant reduction in death or paralysis in mice caused by collagen/epinephrine-induced thromboembolism in a dose-dependent manner and inhibition of collagen-induced human platelet aggregation in a concentration-dependent manner. Additionally, AGE 232-treated mice did not show severe bleeding in the gut compared to ASA-treated mice. AGE 232 resulted in a decrease in the number of neutrophils attached to the human umbilical vein endothelial cells (HUVECs) and lower inhibition of COX-1 in response to bleeding and damage to blood vessels, a major side effect of ASA. Therefore, AGE 232 can prevent thrombus formation and stroke.

The additive memory and healthspan enhancement effects by the combined treatment of mature silkworm powders and Korean angelica extracts.

ETHNOPHARMACOLOGICAL RELEVANCE: Silkworm (Bombyx mori) and Korean angelica (KoAg; Angelica gigas Nakai) have been widely used as traditional oriental medicines in Korea, China, and Japan to treat various diseases such as anemia, cold, diabetes, palsy, stroke, etc. Steamed and freeze-dried mature silkworm powder, also known as HongJam (HJ), and extracts of KoAg root (KoAgE) are currently sold in Korea as functional foods to improve memory, cognition, and liver functions. However, the molecular and pharmacological basis for the improvement of brain functions of HJ and KoAgE has not yet been elucidated. AIM OF STUDY: This study aimed to elucidate the molecular basis underlying the memory-enhancing effects of HJ and KoAgE and determine whether administration of HJ and KoAgE complexes (HJ+KoAgC) has additive memory and healthspan-enhancing effects. MATERIALS AND METHODS: The MCI mouse models generated by intraperitoneal injection of Scopolamine (Sco-IP) were orally administered with HJ and KoAgE alone or as complexes. Their memory-enhancing effects were examined on spatial, fear-aggravated, and social memories and compared with control or Donepezil (Dp) treatment. The activities of mitochondria complex (MitoCom) I-IV and acetylcholinesterase (AChE) and the amounts of ATP in the mouse brains were examined. The Drosophila model was used to investigate lifespan- and healthspan-promoting effects of HJ+KoAgC. RESULTS: Administration of HJ+KoAgC produced more memory-enhancing effects than administration of HJ or KoAgE alone or Dp. The increase in MitoCom I-IV activities and ATP amounts and the decrease in AChE activities in the mouse brains were the molecular basis for the memory enhancement. The greatest improvement in memory and mitochondrial function was observed when the mice were administered the 1:0.8 ratio of HJ+KoAgC. Administration of HJ+KoAgC to Drosophila prolonged the lifespan and the healthspan and increased the amounts of ATP. CONCLUSION: HJ+KoAgC had superior effects on memory improvement and healthspan extension by increasing mitochondrial activities and ATP amounts in treated animal models.