Production of Polyhydroxybutyrate by halotolerant Halomonas cerina YK44 using sugarcane molasses and soybean flour in tap water.

As environmental pollution intensifies, the interest in bioplastics is growing. The bioplastic polyhydroxyalkanoates (PHAs), which are produced and degraded by microorganisms, have received considerable attention. However, the production cost of PHA is still high, and several ways to increase economy of PHA production have been studied. Therefore, as one way of solution, Halomonas species were screened and evaluated with cheap substrates such as molasses and soybean flour. Among tested strains, Halomonas cerina YK44 was selected and used for polyhydroxybutyrate (PHB) production with molasses and soybean flour together, whose combination was not evaluated well before, in tap water. The medium composition optimization showed maximum PHB production at 4 % sugarcane molasses, 2 % NaCl, 0.05 % soybean flour, and pH 8 in tap water (9.2 g/L DCW, 7.3 g/L PHB, and 79.7 % PHB contents). However, cell growth of halotolerant H. cerina YK44 was disturbed by 0.2 % furfural, which existed in biomass based sugars as inhibitors. Physical and thermal analyses revealed that PHB film started from sugarcane molasses and soybean flour was no different from that initiated from simple sugars (Tm was 175.8 °C and 176.2 °C, PDI was 1.29, and 1.31, respectively).

Novel differential scanning calorimetry (DSC) application to select polyhydroxyalkanoate (PHA) producers correlating 3-hydroxyhexanoate (3-HHx) monomer with melting enthalpy.

Polyhydroxyalkanoate (PHA) is an environmental alternative to petroleum-based plastics because of its biodegradability. The polymer properties of PHA have been improved by the incorporation of different monomers. Traditionally, the monomer composition of PHA has been analyzed using gas chromatography (GC) and nuclear magnetic resonance (NMR), providing accurate monomer composition. However, sequential analysis of the thermal properties of PHA using differential scanning calorimetry (DSC) remains necessary, providing crucial insights into its thermal characteristics. To shorten the monomer composition and thermal property analysis, we directly applied DSC to the analysis of the obtained PHA film and observed a high correlation (r2 = 0.98) between melting enthalpy and the 3-hydroxyhexanoate (3-HHx) mole fraction in the polymer. A higher 3-HHx fraction resulted in a lower melting enthalpy as 3-HHx provided the polymer with higher flexibility. Based on this, we selected the poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) (P(3HB-co-3HHx)) producing strain from Cupriavidus strains that newly screened and transformed with vectors containing P(3HB-co-3HHx) biosynthetic genes, achieving an average error rate below 1.8% between GC and DSC results. Cupriavidus sp. BK2 showed a high 3-HHx mole fraction, up to 10.38 mol%, with Tm (℃) = 171.5 and ΔH of Tm (J/g) = 48.0, simultaneously detected via DSC. This study is an example of the expansion of DSC for PHA analysis from polymer science to microbial engineering.

Evaluation of simplified ester-linked fatty acid analysis (ELFA) for phospholipid fatty acid (PLFA) analysis of bacterial population.

Phospholipid fatty acid (PLFA) analysis is used for characterizing microbial communities based on their lipid profiles. This method avoids biases from PCR or culture, allowing data collection in a natural state. However, PLFA is labor-intensive due to lipid fractionation. Simplified ester-linked fatty acid analysis (ELFA), which skips lipid fractionation, offers an alternative. It utilizes base-catalyzed methylation to derivatize only lipids, not free fatty acids, and found glycolipid and neutral lipid fractions are scarcely present in most bacteria, allowing lipid fractionation to be skipped. ELFA method showed a high correlation to PLFA data (r = 0.99) and higher sensitivity than the PLFA method by 1.5-2.57-fold, mainly due to the higher recovery of lipids, which was 1.5-1.9 times higher than with PLFA. The theoretical limit of detection (LOD) and limit of quantification (LOQ) for the ELFA method indicated that 1.54-fold less sample was needed for analysis than with the PLFA method. Our analysis of three bacterial cultures and a simulated consortium revealed the effectiveness of the ELFA method by its simple procedure and enhanced sensitivity for detecting strain-specific markers, which were not detected in PLFA analysis. Overall, this method could be easily used for the population analysis of synthetic consortia.

Dual Application of p-Nitrophenol Alkanoate-Based Assay for Soil Selection and Screening of Microbial Strains for Bioplastic Degradation.

With an increase in the commercialization of bioplastics, the importance of screening for plastic-degrading strains and microbes has emerged. Conventional methods for screening such strains are time-consuming and labor-intensive. Therefore, we suggest a method for quickly and effectively screening plastic-degrading microbial strains through dual esterase assays for soil and isolated strains, using p-nitrophenyl alkanoates as substrates. To select microbe-abundant soil, the total amount of phospholipid fatty acids (PLFAs) included in each soil sample was analyzed, and esterase assays were performed for each soil sample to compare the esterase activity of each soil. In addition, by analyzing the correlation coefficients and sensitivity between the amount of PLFAs and the degree of esterase activity according to the substrate, it was confirmed that substrate pNP-C2 is the most useful index for soil containing several microbes having esterase activity. In addition, esterase assays of the isolated strains allowed us to select the most active strain as the degrading strain, and 16S rRNA results confirmed that it was Bacillus sp. N04 showed the highest degradation activity for polybutylene succinate (PBS) as measured in liquid culture for 7 days, with a degradation yield of 99%. Furthermore, Bacillus sp. N04 showed degradation activity against various bioplastics. We propose the dual application of p-nitrophenyl alkanoates as an efficient method to first select the appropriate soil and then to screen for plastic-degrading strains in it, and conclude that pNP-C2 in particular, is a useful indicator.

Tumor immune microenvironment permissive to metastatic progression of ING4-deficient breast cancer.

Deficiencies in the ING4 tumor suppressor are associated with advanced stage tumors and poor patient survival in cancer. ING4 was shown to inhibit NF-kB in several cancers. As NF-kB is a key mediator of immune response, the ING4/NF-kB axis is likely to manifest in tumor-immune modulation but has not been investigated. To characterize the tumor immune microenvironment associated with ING4-deficient tumors, three approaches were employed in this study: First, tissue microarrays composed of 246 primary breast tumors including 97 ING4-deficient tumors were evaluated for the presence of selective immune markers, CD68, CD4, CD8, and PD-1, using immunohistochemical staining. Second, an immune-competent mouse model of ING4-deficient breast cancer was devised utilizing CRISPR-mediated deletion of Ing4 in a Tp53 deletion-derived mammary tumor cell line; mammary tumors were evaluated for immune markers using flow cytometry. Lastly, the METABRIC gene expression dataset was evaluated for patient survival related to the immune markers associated with Ing4-deleted tumors. The results showed that CD68, CD4, CD8, or PD-1, was not significantly associated with ING4-deficient breast tumors, indicating no enrichment of macrophages, T cells, or exhausted T cell types. In mice, Ing4-deleted mammary tumors had a growth rate comparable to Ing4-intact tumors but showed increased tumor penetrance and metastasis. Immune marker analyses of Ing4-deleted tumors revealed a significant increase in tumor-associated macrophages (Gr-1loCD11b+F4/80+) and a decrease in granzyme B-positive (GzmB+) CD4+ T cells, indicating a suppressive and/or less tumoricidal immune microenvironment. The METABRIC data analyses showed that low expression of GZMB was significantly associated with poor patient survival, as was ING4-low expression, in the basal subtype of breast cancer. Patients with GZMB-low/ING4-low tumors had the worst survival outcomes (HR = 2.80, 95% CI 1.36-5.75, p = 0.0004), supportive of the idea that the GZMB-low immune environment contributes to ING4-deficient tumor progression. Collectively, the study results demonstrate that ING4-deficient tumors harbor a microenvironment that contributes to immune evasion and metastasis.

Identification of Oil-Loving Cupriavidus necator BM3-1 for Polyhydroxyalkanoate Production and Assessing Contribution of Exopolysaccharide for Vegetable Oil Utilization.

Polyhydroxyalkanoates (PHA) have received attention owing to their biodegradability and biocompatibility, with studies exploring PHA-producing bacterial strains. As vegetable oil provides carbon and monomer precursors for poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) (P(3HB-co-3HHx)), oil-utilizing strains may facilitate PHA production. Herein, Cupriavidus necator BM3-1, which produces 11.1 g/L of PHB with 5% vegetable oil, was selected among various novel Cupriavidus necator strains. This strain exhibited higher preference for vegetable oils over sugars, with soybean oil and tryptone determined to be optimal sources for PHA production. BM3-1 produced 33.9 g/L of exopolysaccharides (EPS), which was three-fold higher than the amount produced by H16 (10.1 g/L). EPS exhibited 59.7% of emulsification activity (EI24), higher than that of SDS and of EPS from H16 with soybean oil. To evaluate P(3HB-co-3HHx) production from soybean oil, BM3-1 was engineered with P(3HB-co-3HHx) biosynthetic genes (phaCRa, phaARe, and phaJPa). BM3-1/pPhaCJ produced 3.5 mol% of 3HHx and 37.1 g/L PHA. BM3-1/pCB81 (phaCAJ) produced 32.8 g/L PHA, including 5.9 mol% 3HHx. Physical and thermal analyses revealed that P(3HB-co-5.9 mol% 3HHx) was better than PHB. Collectively, we identified a novel strain with high vegetable oil utilization capacity for the production of EPS, with the option to engineer the strain for P(3HB-co-3HHx).

Controlled production of a polyhydroxyalkanoate (PHA) tetramer containing different mole fraction of 3-hydroxybutyrate (3HB), 3-hydroxyvalerate (3 HV), 4 HV and 5 HV units by engineered Cupriavidus necator.

Polyhydroxyalkanoates (PHAs) are promising alternatives to existing petrochemical-based plastics because of their bio-degradable properties. However, the limited structural diversity of PHAs has hindered their application. In this study, high mole-fractions of Poly (39 mol% 3HB-co-17 mol% 3 HV-co-44 mol% 4 HV) and Poly (25 mol% 3HB-co-75 mol% 5 HV) were produced from 4- hydroxyvaleric acid and 5-hydroxyvaleric acid, using Cupriavidus necator PHB-4 harboring the gene phaCBP-M-CPF4 with modified sequences. In addition, the complex toxicity of precursor mixtures was tested, and it was confirmed that the engineered C. necator was capable of synthesizing Poly (32 mol% 3HB-co-11 mol% 3 HV-co-25 mol% 4 HV-co-32 mol% 5 HV) at low mixture concentrations. Correlation analyses of the precursor ratio and the monomeric mole fractions indicated that each mole fractions could be precisely controlled using the precursor proportion. Physical property analysis confirmed that Poly (3HB-co-3 HV-co-4 HV) is a rubber-like amorphous polymer and Poly (3HB-co-5 HV) has a high tensile strength and elongation at break. Poly (3HB-co-3 HV-co-4 HV-co-5 HV) had a much lower glass transition temperature than the co-, terpolymers containing 3 HV, 4 HV and 5 HV. This study expands the range of possible physical properties of PHAs and contributes to the realization of custom PHA production by suggesting a method for producing PHAs with various physical properties through mole-fraction control of 3 HV, 4 HV and 5 HV.

Establishment of efficient 5-hydroxyvaleric acid production system by regenerating alpha-ketoglutaric acid and its application in poly(5-hydroxyvaleric acid) production.

5-hydroxyvaleric acid (5-HV) is a versatile C5 intermediate of bio-based high-value chemical synthesis pathways. However, 5-HV production faces a few shortcomings involving the supply of cofactors, especially α-ketoglutaric acid (α-KG). Herein, we established a two-cell biotransformation system by introducing L-glutamate oxidase (GOX) to regenerate α-KG. Additionally, the catalase KatE was adapted to inhibit α-KG degradation by the H2O2 produced during GOX reaction. We searched for the best combination of genes and vectors and optimized the biotransformation conditions to maximize GOX effectiveness. Under the optimized conditions, 5-HV pathway with GOX showed 1.60-fold higher productivity than that of without GOX, showing 11.3 g/L titer. Further, the two-cell system with GOX and KatE was expanded to produce poly(5-hydroxyvaleric acid) (P(5HV)), and it reached at 412 mg/L of P(5HV) production and 20.5% PHA contents when using the biotransformation supernatant. Thus, the two-cell biotransformation system with GOX can potentially give the practical and economic alternative of 5-HV production using bio-based methods. We also propose direct utilization of 5-HV from bioconversion for P(5HV) production.

Reproducible Polybutylene Succinate (PBS)-Degrading Artificial Consortia by Introducing the Least Type of PBS-Degrading Strains.

Polybutylene succinate (PBS) stands out as a promising biodegradable polymer, drawing attention for its potential as an eco-friendly alternative to traditional plastics due to its biodegradability and reduced environmental impact. In this study, we aimed to enhance PBS degradation by examining artificial consortia composed of bacterial strains. Specifically, Terribacillus sp. JY49, Bacillus sp. JY35, and Bacillus sp. NR4 were assessed for their capabilities and synergistic effects in PBS degradation. When only two types of strains, Bacillus sp. JY35 and Bacillus sp. NR4, were co-cultured as a consortium, a notable increase in degradation activity toward PBS was observed compared to their activities alone. The consortium of Bacillus sp. JY35 and Bacillus sp. NR4 demonstrated a remarkable degradation yield of 76.5% in PBS after 10 days. The degradation of PBS by the consortium was validated and our findings underscore the potential for enhancing PBS degradation and the possibility of fast degradation by forming artificial consortia, leveraging the synergy between strains with limited PBS degradation activity. Furthermore, this study demonstrated that utilizing only two types of strains in the consortium facilitates easy control and provides reproducible results. This approach mitigates the risk of losing activity and reproducibility issues often associated with natural consortia.

5-aminosalicylic acid suppresses osteoarthritis through the OSCAR-PPARγ axis.

Osteoarthritis (OA) is a progressive and irreversible degenerative joint disease that is characterized by cartilage destruction, osteophyte formation, subchondral bone remodeling, and synovitis. Despite affecting millions of patients, effective and safe disease-modifying osteoarthritis drugs are lacking. Here we reveal an unexpected role for the small molecule 5-aminosalicylic acid (5-ASA), which is used as an anti-inflammatory drug in ulcerative colitis. We show that 5-ASA competes with extracellular-matrix collagen-II to bind to osteoclast-associated receptor (OSCAR) on chondrocytes. Intra-articular 5-ASA injections ameliorate OA generated by surgery-induced medial-meniscus destabilization in male mice. Significantly, this effect is also observed when 5-ASA was administered well after OA onset. Moreover, mice with DMM-induced OA that are treated with 5-ASA at weeks 8-11 and sacrificed at week 12 have thicker cartilage than untreated mice that were sacrificed at week 8. Mechanistically, 5-ASA reverses OSCAR-mediated transcriptional repression of PPARγ in articular chondrocytes, thereby suppressing COX-2-related inflammation. It also improves chondrogenesis, strongly downregulates ECM catabolism, and promotes ECM anabolism. Our results suggest that 5-ASA could serve as a DMOAD.

Enhanced theanine production with reduced ATP supply by alginate entrapped Escherichia coli co-expressing γ-glutamylmethylamide synthetase and polyphosphate kinase.

L-theanine is an amino acid with a unique flavor and many therapeutic effects. Its enzymatic synthesis has been actively studied and γ-Glutamylmethylamide synthetase (GMAS) is one of the promising enzymes in the biological synthesis of theanine. However, the theanine biosynthetic pathway with GMAS is highly ATP-dependent and the supply of external ATP was needed to achieve high concentration of theanine production. As a result, this study aimed to investigate polyphosphate kinase 2 (PPK2) as ATP regeneration system with hexametaphosphate. Furthermore, the alginate entrapment method was employed to immobilize whole cells containing both gmas and ppk2 together resulting in enhanced reusability of the theanine production system with reduced supply of ATP. After immobilization, theanine production was increased to 239 mM (41.6 g/L) with a conversion rate of 79.7% using 15 mM ATP and the reusability was enhanced, maintaining a 100% conversion rate up to the fifth cycles and 60% of conversion up to eighth cycles. It could increase long-term storage property for future uses up to 35 days with 75% activity of initial activity. Overall, immobilization of both production and cofactor regeneration system could increase the stability and reusability of theanine production system.

Positive effect of phasin in biohydrogen production of non polyhydroxybutyrate-producing Clostridium acetobutylicum ATCC 824.

In this study, the goal was to enhance the tolerance of Clostridium acetobutylicum ATCC 824 to biomass-based inhibitory compounds for biohydrogen production and evaluate various known genes that enhance the production of biochemicals in various hosts. The introduction of phaP, the major polyhydroxyalkanoate granule-associated protein that has been reported as a chaperone-like protein resulted in increased tolerance to inhibitors and leads to higher levels of hydrogen production, cell growth, and glucose consumption in the presence of these inhibitors. It was observed that the introduction of phaP led to an increase in the transcription of the hydrogenase gene, whereas transcription of the chaperone functional genes decreased compared to the wild type. Finally, the introduction of phaP could significantly enhance biohydrogen production by 2.6-fold from lignocellulosic hydrolysates compared to that of wild type. These findings suggested that the introduction of phaP could enhance growth and biohydrogen production, even in non-polyhydroxyalkanoate-producing strains.

Prevalence of porcine circovirus type 2 and type 3 in slaughtered pigs and wild boars in Korea.

BACKGROUND: Porcine circovirus, a non-enveloped single-stranded DNA virus belonging to the genus Circovirus of the family Circoviridae, is a major pathogen of porcine circovirus-associated disease. Porcine circovirus 3, a novel porcine circovirus, has been identified in individuals with clinical symptoms. OBJECTIVES: The prevalence of porcine circovirus 2 and porcine circovirus 3 and the confirmation of diagnosis of this emerging viral disease have not been fully studied yet. Therefore, the objective of the present study was to investigate the prevalence of porcine circovirus 2 and porcine circovirus 3 in slaughtered pigs and wild boars in Korea between 2018 and 2019. METHODS: Lungs and hilar lymph nodes of healthy pigs slaughtered in slaughterhouses and captured wild pigs were collected, and viruses were detected by multiplex quantitative polymerase chain reaction and two staining methods (in situ hybridization and immunohistochemistry) to confirm the presence of porcine circovirus 2 and porcine circovirus 3. RESULTS: Positive rates of porcine circovirus 2 in lungs and hilar lymph nodes were 78.1% (75/96) and 89.5% (86/96) in slaughtered pigs, respectively. They were 18.0% (30/167) and 46.3% (24/55) in wild boars, respectively. Positive rates of porcine circovirus 3 in lungs and hilar lymph nodes were 30.2% (29/96) and 13.5% (13/96) in slaughtered pigs, respectively. They were 4.2% (7/167) and 5.5% (3/55) in wild boars, respectively. At the farm level, positive rates of porcine circovirus 2 and porcine circovirus 3 were 97.9% (47/48) and 54.2% (26/48), respectively. Positive rates of porcine circovirus 2 and porcine circovirus 3 decreased in spring. Immunohistochemistry and in situ hybridization confirmed the presence of porcine circovirus 2 and porcine circovirus 3 in lungs, but not porcine circovirus 3 in the hilar lymph nodes. CONCLUSION: These results suggest that the prevalence of porcine circovirus 2 and porcine circovirus 3 might vary depending on the season and the type of sample. Wild boars might play a role in the epidemiology of porcine circovirus 2 and porcine circovirus 3 in South Korea. Continuous surveillance and further study are needed for this emerging disease.

Amentoflavone, a potent natural matrix metalloproteinase 2 inhibitor.

Gelatinase A (MMP-2) has been studied and proven to play a vital role in the intrusion and metastasis of cancer. Flavonoids influence on molecular and cellular functions of MMP-2 and thus a systematic investigation of flavonoids against the metalloproteolytic activity of MMP-2 has been performed in this study. A fluorescence resonance energy transfer method was used to investigate the inhibitory activities of various flavonoids. Flavone, flavonol and isobavachalcone derivatives showed their inhibitory activity against MMP-2. Surprisingly, the most effective inhibitor was Amentoflavone and its blocking function was superior to other flavonoids. Its IC50 value was 0.689 µM. An induced-fit docking study was carried out to survey its extraordinary activity. The binding mode of Amentoflavone is quite similar to that of (2 ∼ {S})-2-[2-[4-(4-methoxyphenyl) phenyl] sulfanylphenyl] pentanedioic acid complexed with MMP-9. Amentoflavone interacts with the functional zinc and catalytic residue, Glu202. Therefore, the docking study reasonably confirmed the strong inhibitory activity of Amentoflavone.

A Study of Drug Repurposing to Identify SARS-CoV-2 Main Protease (3CLpro) Inhibitors.

The outbreak of coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) wreaked havoc all over the world. Although vaccines for the disease have recently become available and started to be administered to the population in various countries, there is still a strong and urgent need for treatments to cure COVID-19. One of the safest and fastest strategies is represented by drug repurposing (DRPx). In this study, thirty compounds with known safety profiles were identified from a chemical library of Phase II-and-up compounds through a combination of SOM Biotech's Artificial Intelligence (AI) technology, SOMAIPRO, and in silico docking calculations with third-party software. The selected compounds were then tested in vitro for inhibitory activity against SARS-CoV-2 main protease (3CLpro or Mpro). Of the thirty compounds, three (cynarine, eravacycline, and prexasertib) displayed strong inhibitory activity against SARS-CoV-2 3CLpro. VeroE6 cells infected with SARS-CoV-2 were used to find the cell protection capability of each candidate. Among the three compounds, only eravacycline showed potential antiviral activities with no significant cytotoxicity. A further study is planned for pre-clinical trials.

Cxcl10 Chemokine Induces Migration of ING4-Deficient Breast Cancer Cells via a Novel Cross Talk Mechanism between the Cxcr3 and Egfr Receptors.

The chemokine Cxcl10 has been associated with poor prognosis in breast cancer, but the mechanism is not well understood. Our previous study has shown that CXCL10 was repressed by the ING4 tumor suppressor, suggesting a potential inverse functional relationship. We thus investigated a role for Cxcl10 in the context of ING4 deficiencies in breast cancer. We first analyzed public gene expression data sets and found that patients with CXCL10-high/ING4-low expressing tumors had significantly reduced disease-free survival in breast cancer. In vitro, Cxcl10 induced migration of ING4-deleted breast cancer cells but not of ING4-intact cells. Using inhibitors, we found that Cxcl10-induced migration of ING4-deleted cells required Cxcr3, Egfr, and the Gßγ subunits downstream of Cxcr3 but not Gαi. Immunofluorescent imaging showed that Cxcl10 induced early transient colocalization between Cxcr3 and Egfr in both ING4-intact and ING4-deleted cells, which recurred only in ING4-deleted cells. A peptide agent that binds to the internal juxtamembrane domain of Egfr inhibited Cxcr3/Egfr colocalization and cell migration. Taken together, these results presented a novel mechanism of Cxcl10 that elicits migration of ING4-deleted cells, in part by inducing a physical or proximal association between Cxcr3 and Egfr and signaling downstream via Gßγ. These results further indicated that ING4 plays a critical role in the regulation of Cxcl10 signaling that enables breast cancer progression.

A Study of 3CLpros as Promising Targets against SARS-CoV and SARS-CoV-2.

The outbreak of coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), results in serious chaos all over the world. In addition to the available vaccines, the development of treatments to cure COVID-19 should be done quickly. One of the fastest strategies is to use a drug-repurposing approach. To provide COVID-19 patients with useful information about medicines currently being used in clinical trials, twenty-four compounds, including antiviral agents, were selected and assayed. These compounds were applied to verify the inhibitory activity for the protein function of 3CLpros (main proteases) of SARS-CoV and SARS-CoV-2. Among them, viral reverse-transcriptase inhibitors abacavir and tenofovir revealed a good inhibitory effect on both 3CLpros. Intriguingly, sildenafil, a cGMP-specific phosphodiesterase type 5 inhibitor also showed significant inhibitory function against them. The in silico docking study suggests that the active-site residues located in the S1 and S2 sites play key roles in the interactions with the inhibitors. The result indicates that 3CLpros are promising targets to cope with SAR-CoV-2 and its variants. The information can be helpful to design treatments to cure patients with COVID-19.

A study of inhibitors of d-glycero-ß-d-manno-heptose-1-phosphate adenylyltransferase from Burkholderia pseudomallei as a potential antibiotic target.

d-Glycero-ß-d-manno-heptose-1-phosphate adenylyltransferase from Burkholderia pseudomallei (BpHldC) is the fourth enzyme in the ADP-l-glycero-ß-d-manno-heptose biosynthesis pathway producing a lipopolysaccharide core. Therefore, BpHldC is an anti-melioidosis target. Three ChemBridge compounds purchased from ChemBridge Corporation (San Diego, CA) were found to have an effective inhibitory activity on BpHldC. Interestingly, ChemBridge 7929959 was the most effective compound due to the presence of the terminal benzyl group. The enzyme kinetic study revealed that most of them show mixed type inhibitory modes against ATP and ßG1P. The induced-fit docking indicated that the medium affinity of ChemBridge 7929959 is originated from its benzyl group occupying the substrate-binding pocket of BpHldC. The inhibitory role of terminal aromatic groups was proven with ChemBridge 7570508. Combined with the previous study, ChemBridge 7929959 is found to work as a dual inhibitor against both HldC and HddC. Therefore, three ChemBridge compounds can be developed as a potent anti-melioidosis agent with a novel inhibitory concept.

Efficient 0.75-mJ 100-kHz ultraviolet pulsed laser based on a rod-type photonic crystal fiber.

We have developed a high-power ultraviolet (UV) nanosecond-pulsed laser based on a rod-type photonic crystal fiber. The UV pulse energy and the pulse repetition rate are 0.75 mJ and 100 kHz, respectively, yielding 75-W UV average power. The temporal pulse shape and the linewidth of a 1030-nm seed laser are optimized for efficient third-harmonic generation, and the high conversion efficiency of 50% is achieved with a good beam quality (M2∼1.2). To our knowledge, this is the highest UV pulse energy from fiber lasers.

A triple-targeting inhibitory activity of Rose Bengal on polysaccharide biosynthesis of Burkholderia pseudomallei.

Sugar nucleotidyltransferases (SNTs) participate in various biosynthesis pathways constructing polysaccharides in Gram-negative bacteria. In this study, a triple-targeting inhibitory activity of Rose Bengal against SNTs such as d-glycero-α-d-manno-heptose-1-phosphate guanylyltransferase (HddC), d-glycero-ß-d-manno-heptose-1-phosphate adenylyltransferase (HldC), and 3-deoxy-d-manno-oct-2-ulosonic acid cytidylyltransferase (KdsB) from Burkholderia pseudomallei is provided. Rose Bengal effectively suppresses the nucleotidyltransferase activity of the three SNTs, and its IC50 values are 10.42, 0.76, and 5.31 µM, respectively. Interestingly, Rose Bengal inhibits the three enzymes regardless of their primary, secondary, tertiary, and quaternary structural differences. The experimental results indicate that Rose Bengal possesses the plasticity to shape its conformation suitable to interact with the three SNTs. As HddC functions in the formation of capsular polysaccharides and HldC and KdsB produce building blocks to constitute the inner core of lipopolysaccharide, Rose Bengal is a potential candidate to design antibiotics in a new category. In particular, it can be developed as a specific antimelioidosis agent. As the mortality rate of the infected people caused by B. pseudomallei is quite high, there is an urgent need for specific antimelioidosis agents. Therefore, a further study is being carried out with derivatives of Rose Bengal.