Effects of Alginic Acid on the Porcine Granulosa Cells and Maturation of Porcine Oocytes.

SCOPE: Alginic acid (AA) from brown algae is a marine organic compound. There is extensive use of AA in the food industry and healthcare, suggesting a high probability of AA exposure. The present study investigates the effects of AA on porcine ovarian granulosa cells (GCs) and oocytes to explore its mechanism in female reproduction because of its adverse effects on reproduction. METHODS AND RESULTS: The study adds 20 µM AA to the porcine primary ovarian GCs medium and porcine oocyte in vitro maturation (IVM) medium. Estrogen and progesterone levels are downregulated in GCs. Reactive oxygen species are excessive, and the antioxidant capacity declines. Then mitochondria-mediated apoptosis pathway is involved in GCs apoptosis. In addition, scores of autophagosomes are found in the experimental cells. Furthermore, AA significantly inhibits the proliferation of GCs around cumulus-oocyte complexes (COCs) accompanied by abnormal spindle assembly, chromosome arrangement disorder, and aberrant cortical granules distribution in oocytes, leading to a decreased oocyte maturation rate. CONCLUSION: These findings suggest that 20 µM AA is toxic to sow reproduction by interfering with estrogen production, oxidative stress, mitochondria-mediated apoptosis, autophagy in GCs of sows, and oocyte maturation.

An Oligomannuronic Acid-Sialic Acid Conjugate Capable of Inhibiting Aß42 Aggregation and Alleviating the Inflammatory Response of BV-2 Microglia.

Oligomannuronic acid (MOS) from seaweed has antioxidant and anti-inflammatory activities. In this study, MOS was activated at the terminal to obtain three different graft complexes modified with sialic acid moiety (MOS-Sia). The results show that MOS-Sia addition can reduce the ß-structure formation of Aß42, and the binding effect of MOS-Sia3 is more obvious. MOS-Sia conjugates also have a better complexing effect with Ca2+ while reducing the formation of Aß42 oligomers in solutions. MOS-Sia3 (25-50 µg/mL) can effectively inhibit the activation state of BV-2 cells stimulated by Aß42, whereas a higher dose of MOS-Sia3 (>50 µg/mL) can inhibit the proliferation of BV-2 cells to a certain extent. A lower dose of MOS-Sia3 can also inhibit the expression of IL-1ß, IL-6, TNF-α, and other proinflammatory factors in BV-2 cells induced by Aß42 activation. In the future, the MOS-Sia3 conjugate can be used to treat Alzheimer's disease.

Integrated regulatory network in Pseudomonas syringae reveals dynamics of virulence.

Pseudomonas syringae, a Gram-negative plant pathogen, expresses multitudinous transcriptional regulators to control the type III secretion system (T3SS) and response to diverse environmental challenges. Although the mechanisms of virulence-associated regulators of P. syringae have been studied for decades, the overall crosstalk underlying these regulators is still elusive. Here, we identify five T3SS regulators (EnvZ-OmpR, CbrAB2, PhoPQ, PilRS, and MgrA), and find that the two-component systems EnvZ-OmpR and CbrAB2 negatively regulate the T3SS. To elucidate crosstalk between 16 virulence-associated regulators in P. syringae, we map an online intricate network called "PSRnet" (Pseudomonas syringae regulatory network) by combining the differentially expressed genes (DEGs) of these 16 regulators by RNA sequencing (RNA-seq) and their binding loci by chromatin immunoprecipitation sequencing (ChIP-seq). Consequently, we identify 238 and 153 functional genes involved in the T3SS and other virulence-related pathways in KB and MM media, respectively. Our results provide insights into the mechanism of plant infections caused by P. syringae.

Structural and biochemical characterisation of a novel alginate lyase from Paenibacillus sp. str. FPU-7.

A novel alginate lyase, PsAly, with a molecular mass of 33 kDa and whose amino acid sequence shares no significant similarity to other known proteins, was biochemically and structurally characterised from Paenibacillus sp. str. FPU-7. The maximum PsAly activity was obtained at 65 °C, with an optimum pH of pH 7-7.5. The activity was enhanced by divalent cations, such as Mg2+, Mn2+, or Co2+, and inhibited by a metal chelator, ethylenediaminetetraacetic acid. The reaction products indicated that PsAly is an endolytic enzyme with a preference for polymannuronate. Herein, we report a detailed crystal structure of PsAly at a resolution of 0.89 Å, which possesses a ß-helix fold that creates a long cleft. The catalytic site was different from that of other polysaccharide lyases. Site-directed mutational analysis of conserved residues predicted Tyr184 and Lys221 as catalytic residues, abstracting from the C5 proton and providing a proton to the glycoside bond, respectively. One cation was found to bind to the bottom of the cleft and neutralise the carboxy group of the substrate, decreasing the pKa of the C5 proton to promote catalysis. Our study provides an insight into the structural basis for the catalysis of alginate lyases and ß-helix polysaccharide lyases.

Characterization of a bifunctional alginate lyase as a new member of the polysaccharide lyase family 17 from a marine strain BP-2.

OBJECTIVES: Bifunctional alginate lyase can efficiently saccharify alginate biomass and prepare functional oligosaccharides of alginate. RESULTS: A new BP-2 strain that produces alginate lyase was screened and identified from rotted Sargassum. A new alginate lyase, Alg17B, belonging to the polysaccharide lyase family 17, was isolated and purified from BP-2 fermentation broth by freeze-drying, dialysis, and ion exchange chromatography. The enzymatic properties of the purified lyase were investigated. The molecular weight of Alg17B was approximately 77 kDa, its optimum reaction temperature was 40-45 °C, and its optimum reaction pH was 7.5-8.0. The enzyme was relatively stable at pH 7.0-8.0, with a temperature range of 25-35 °C, and the specific activity of the purified enzyme reached 4036 U/mg. A low Na+ concentration stimulated Alg17B enzyme activity, but Ca2+, Zn2+, and other metal ions inhibited it. Substrate specificity analysis, thin-layer chromatography, and mass spectrometry showed that Alg17B is an alginate lyase that catalyses the hydrolysis of sodium alginate, polymannuronic acid (polyM) and polyguluronic acid to produce monosaccharides and low molecular weight oligosaccharides. Alg17B is also bifunctional, exhibiting both endolytic and exolytic activities toward alginate, and has a wide substrate utilization range with a preference for polyM. CONCLUSIONS: Alg17B can be used to saccharify the main carbohydrate, alginate, in the ethanolic production of brown algae fuel as well as in preparing and researching oligosaccharides.

Genetic, structural and pharmacological characterization of polymannuronate synthesized by algG mutant indigenous soil bacterium Pseudomonas aeruginosa CMG1421.

AIMS: It was aimed to study the genetic, structural and pharmacological characteristics of polymannuronate synthesized by Pseudomonas aeruginosa CMG1421. METHODS AND RESULTS: Synthesis was analysed by transmission electron microscopy, FT/IR, 1 H-NMR and gel permeation chromatography followed by in vitro bioassays. Colony PCR followed by sequence analysis was employed for screening of structural genes. FT/IR analysis indicated the presence of hydroxyl, carboxyl and O-acetyl groups linked to mannuronate. 1 H-NMR analysis indicated M-M bond characteristics for mannuronic acid residues. The average relative molecular weight was found in range of 20 000-250 000 Da. The amplified DNA fragments were identified as 16S rRNA, algD, alg8, alg44, algG, algE and algX genes showing 99-100% homology with those of P. aeruginosa. However, in algG there were transition mutations of adenine and cytosine at nucleotide position 766 and 769, and 878 and 881 respectively. Polymannuronate and its oligomannuronates respectively showed moderate and significant antioxidant, anti-inflammatory, anti-obesity and antidiabetic activities. CONCLUSIONS: Alginate synthesized by ∆algG mutant P. aeruginosa CMG1421 was bioactive and solely consists of acetylated d-mannuronates. SIGNIFICANCE AND IMPACT OF THE STUDY: We investigated biocompatible, nonimmunogenic and nontoxic pharmacological agents for treatment and attenuation of degenerative, inflammatory, autoimmune disease, and metabolic disorders such as obesity and diabetes.

Upgrading the antioxidant properties of fucoidan and alginate from Cystoseira trinodis by fungal fermentation or enzymatic pretreatment of the seaweed biomass.

The seaweed Cystoseira trinodis was fermented by different fungi prior to extraction of fucoidan and alginate to enhance their antioxidative potential. All the investigated fungi were able to produce fucoidanase (1.05-3.41 U/ml) and alginate lyase (7.27-18.59 U/mL). Different fungal species induced a reduction in the molecular weight (MW) of fucoidan and alginate in comparison to the unfermented control. The MW of fucoidan reduced by 41-81.5%, while the MW of alginate was reduced by 28-75%, depending on the fungal species. Significant increases in the fucose and sulphate contents of fucoidan and mannuronic/guluronic acid ratio of alginate were induced by fungal fermentation. Fungal pretreatment enhanced the ferric reducing antioxidant power, total antioxidant capacity and hydroxyl radical scavenging activity of both fucoidan and alginate. Additionally, enzymatic pretreatment of the macroalgal biomass assisted in the recovery of fucoidan and alginate with low molecular weight and enhanced antioxidative potential.

Multi-Drug Resistant Clinical Pseudomonas aeruginosas Inhibited by Ferula gummosa Boiss.

BACKGROUND: Multi-drug resistance among Pseudomonas aeruginosa (P. aeruginosa) clinical isolates is increasing and becoming a serious problem for public health authorities worldwide. OBJECTIVE: The aim of the current study is to introduce a potent antibacterial compound against the resistant P. aeruginosa. METHODS: In this study, we evaluated the antibacterial effects of extracts and essential oils of Ferula gummosa Boiss (F. gummosa) on 33 P. aeruginosa clinical isolates by microdilution method and assessed the association of antimicrobial activity with the extended spectrum ß-lactamase (ESBL) producing, biofilm forming and aliginate production of the strains. In addition, the presence of some genes involved in these properties, including blaGES- 1, blaRER-1, blaCTX-M, blaVEB-1, blaOXA-1, blaOXA-4, blaOXA-10, ppyR, pslA, pelA, algU, algL, algD, fliC and oxaA was determined using PCR. RESULTS: We revealed that all of our extracts and essential oils had significant antibacterial effects (p<0.001), but the aqueous extracts showed a relatively lower antibacterial activity compared with the methanolic ones. Furthermore, the minimum inhibitory concentration required for the ESBL producing strains was significantly higher than the non-ESBL producing ones (p<0.001). Loss of some genes such as blaPER-1, blaGES-1, blaOXA-1 and blaOXA-4 caused sensitivity to F. gummosa derivatives (p<0.05). CONCLUSION: The findings of this study indicate that the antibacterial effects of the extract and essential oils of F. gummosa may be a potential novel treatment against drug-resistant P. aeruginosa clinical isolates.