A calorimetric study of thermal denaturation of recombinant human lactoferrin from rice

Thermal denaturation of recombinant human lactoferrin from transgenic rice with different degrees of iron saturation has been studied by differential scanning calorimetry. The maximum temperature, enthalpy change, and activation energy of denaturation were higher when recombinant lactoferrin was more saturated with iron, indicating an increase in the stability of the protein structure. Maximum temperature and activation energy values for apo- and holo-lactoferrins were practically identical to those reported for the same forms of lactoferrin from human milk, which indicates a similar thermal stability. However, the value of enthalpy change for denaturation of the recombinant lactoferrin was 2.5-3-fold lower than that found for the human milk protein. This finding may reflect the influence that the different glycosylation pattern may have in the relationship between lactoferrin domains. Denaturation of recombinant lactoferrin in milk was compared with denaturation in phosphate buffer, and results indicated that the protein was more heat-sensitive when treated in milk than in buffer.     

Broccoli processing wastes as a source of peroxidase

A peroxidase isozyme (BP) was purified to homogeneity from broccoli stems ( Brassica oleraceae var. maraton) discarded from industrial processing wastes. BP specific activity was 1216 ABTS [2,2'-azinobis(3-ethylbenzthiazoline-6-sulfonic acid)] units/mg, representing 466-fold that of crude extract. BP is a monomeric glycoprotein containing 16% carbohydrates, with a molecular mass of 49 kDa and an isoelectric point close to 4.2. From kinetic data it showed a two-substrate ping-pong mechanism, and the catalytic efficiency measured as the rate-limiting step of free BP regeneration was 3.4 x 10(6) M(-1) s(-1). The ABTS K m value was 0.2 mM, which was about 20 times lower than that reported for acidic commercial horseradish peroxidase (HRP). Assessment of BP secondary structure showed 30% helical character, similar to HRP and cytochrome c peroxidase. BP lost only 25% activity after 10 min of heating at 55 degrees C and pH 6; it was stable in the pH range from 4 to 9 and showed an optimum pH of 4.6 using ABTS as substrate. BP was active on substrates normally involved in lignin biosynthesis, such as caffeic and ferulic acids, and also displayed good catechol oxidation activity in the presence of hydrogen peroxide. Reverse micellar extraction was successfully used as potential large-scale prepurification of broccoli peroxidase, achieving a purification factor of 7, with 60% activity yield. Stems from the broccoli processing industry have a high potential as an alternative for peroxidase purification.     

Functional CpG methylation system in a social insect

DNA methylation systems are well characterized in vertebrates, but methylation in Drosophila melanogaster and other invertebrates remains controversial. Using the recently sequenced honey bee genome, we present a bioinformatic, molecular, and biochemical characterization of a functional DNA methylation system in an insect. We report on catalytically active orthologs of the vertebrate DNA methyltransferases Dnmt1 and Dnmt3a and b, two isoforms that contain a methyl-DNA binding domain, genomic 5-methyl-deoxycytosine, and CpG-methylated genes. The honey bee provides an opportunity to study the roles of methylation in social contexts.     

Updated Trends on the Biodiscovery of New Marine Natural Products from Invertebrates

From 1990–2019, a total of 15,442 New Marine Natural Products from Invertebrates (NMNPIs) were reported. The 2010s saw the most prolific decade of biodiscovery, with 5630 NMNPIs recorded. The phyla that contributed most biomolecules were the Porifera (sponges) (47.2%, 2659 NMNPIs) and the Cnidaria (35.3%, 1989 NMNPIs). The prevalence of these two phyla as the main sources of NMNPIs became more pronounced in the 2010s. The tropical areas of the Pacific Ocean yielded more NMNPIs, most likely due to the remarkable biodiversity of coral reefs. The Indo-Burma biodiversity hotspot (BH) was the most relevant area for the biodiscovery of NMNPIs in the 2010s, accounting for nearly one-third (1819 NMNPIs) of the total and surpassing the top BH from the 1990s and the 2000s (the Sea of Japan and the Caribbean Islands, respectively). The Chinese exclusive economic zone (EEZ) alone contributed nearly one-quarter (24.7%) of all NMNPIs recorded during the 2010s, displacing Japan’s leading role from the 1990s and the 2000s. With the biodiscovery of these biomolecules steadily decreasing since 2012, it is uncertain whether this decline has been caused by lower bioprospecting efforts or the potential exhaustion of chemodiversity from traditional marine invertebrate sources.     

The cell and molecular basis of mechanical, cold, and inflammatory pain

Peripheral pain pathways are activated by a range of stimuli. We used diphtheria toxin to kill all mouse postmitotic sensory neurons expressing the sodium channel Nav1.8. Mice showed normal motor activity and low-threshold mechanical and acute noxious heat responses but did not respond to noxious mechanical pressure or cold. They also showed a loss of enhanced pain responses and spontaneous pain behavior upon treatment with inflammatory insults. In contrast, nerve injury led to heightened pain sensitivity to thermal and mechanical stimuli indistinguishable from that seen with normal littermates. Pain behavior correlates well with central input from sensory neurons measured electrophysiologically in vivo. These data demonstrate that Na(v)1.8-expressing neurons are essential for mechanical, cold, and inflammatory pain but not for neuropathic pain or heat sensing.     

Accuracy of pairwise methods in the reconstruction of family relationships, using molecular information from turbot (Scophthalmus maximus)

Many estimators and algorithms have been developed to infer the genealogical relationships from molecular marker data when there is no pedigree information. Most pairwise methods provide estimates in a continuous range that needs to be converted into genealogical relationships (namely full-sibs, half-sibs and unrelated) if there is a previous knowledge of the population structure. Transformations are usually based on arbitrary thresholds, but the possibility of correcting the coancestry estimates via explicit pedigree reconstruction methods has been suggested. Using molecular data for ten highly polymorphic microsatellite loci on a population of turbot (Scophthalmus maximus) with a known genealogy, the behaviour of four pairwise marker-based coancestry estimators and the molecular coancestry has been evaluated. The population consisted on 138 families with 4 full-sib individuals each and one family with 8 full-sib individuals (up to 15 half-sib families were also present due to the sharing of parents between some of the full-sibs families). Our results suggested that transforming the pairwise estimators and the molecular coancestry to family relationships through the establishment of thresholds performs slightly better than the explicit pedigree reconstruction method, when accuracy is measured in a pairwise basis. However, if joint relationships between more than two individuals were tested at a time, the threshold methods led to a high percentage of incongruous triads of full-sib individuals, with Mendelian incompatibilities appearing in congruous full-sib families (more than 70% and 60% in our study, respectively). The explicit pedigree reconstruction approach, due to its nature, is free from such problems. Therefore, the pedigree reconstruction approach seems to be a valuable tool to provide a congruent and compatible pedigree when the pairwise marker-based coancestry matrices or the molecular coancestry need to be transformed.     

Effect of different methods for the induction of spermiation on semen quality in European eel

Five hormonal treatments with human chorionic gonadotropin (hCG) were tested for the induction of maturation and spermiation in male farmed eels. The main aim was to optimize previously used hormonal treatments to achieve shorter induction treatments, longer spermiation periods and/or higher sperm quality. Fish treated for just 3 weeks (treatment E) or until the onset of spermiation (treatment C) showed the worst results, while the treatment consisting of weekly administration of 1.5 IU hCG g^?1 fish (treatment A) induced the highest percentage of spermiating males, the highest number of sperm samples and sperm volumes and densities similar to the rest of the treatments (B: half hormone dosage, or D: biweekly administration). Evaluation of the sperm quality was performed by computer‐assisted sperm analysis (CASA), considering the percentage of total motile spermatozoa, the percentage of fast and medium‐velocity spermatozoa, as well as different motility parameters. Sperm samples from A‐D groups showed between 44% and 54% motile spermatozoa, and between 10% and 15% fast spermatozoa, while samples from E‐treated males showed 0% motile cells. No significant differences were found in the spermatozoa straight line velocity (VSL), curvilinear velocity (VCL) or the angular velocity (VAP), neither spermatozoa beating cross frequency (BCF) between A–D groups.     

Effects of Vermicompost and Vermiwash on Plant,Phenolic Content,and Anti-oxidant Activity of Mexican Pepperleaf (Piper auritum Kunth) Cultivated in Phosphate Rock Potting Media

The aim of this investigation was to determine the effect of vermicompost, vermiwash, and phosphate rock on plant, total phenols, flavonoids, and anti-oxidant activity in Piper auritum Kunth leaves. P. auritum plants were obtained from cuttings and were planted according to the Box-Behnken experimental design with three repetitions at the central point. The factors and levels were vermicompost (10, 20, and 30 g plant^?1), vermiwash (5, 10, and 15 mL plant^?1), and phosphate rock (1, 2, and 3 g plant^?1). Plant growth parameters (plant height, stem diameter, leaves number) and chlorophyll content were measured 1 month after treatment applications. Total phenols, total flavonoids, and 1,1-diphenyl-2-picryl-hydrazyl radical scavenging activity was measured after 4 months. Vermicompost, vermiwash, and phosphate rock had no statistically significant effect on plant growth. Plant height, stem diameter, leaves number, chlorophyll, innermost number, fresh weight stem, fresh weight leaves, fresh weight root, dry weight stem, dry weight leaves, and dry weight root were not different among treatments. Total phenolic compounds were statistically affected for both vermiwash and phosphoric rock (p < 0.05) and the anti-oxidant activity decreased by vermicompost addition. The application of 15 mL plant^?1 vermiwash, 1 g phosphate rock, and 20 g vermicompost plant^?1 increased the total phenol content.     

Effect of hypo- and hyper-saline conditions on osmolarity and fatty acid composition of juvenile shrimp Litopenaeus vannamei (Boone, 1931) fed low- and high-HUFA diets

Litopenaeus vannamei (Boone) grown in ponds are exposed to salinities of less than 5 g L^?1 during inland shrimp culture or to more than 40 g L^?1 from evaporation and reduced water exchange in dry, hot climates. However, dietary requirements for shrimp grown in low or high salinities are not well defined, particularly for fatty acids. Feeding shrimp postlarvae with highly unsaturated fatty acids (HUFA) enhances tolerance to acute exposure to low salinity, as a result of better nutritional status, or/and specific effects of HUFA on membrane function and osmoregulation mechanisms. This study analysed the effect of HUFA supplementation (3% vs. 34%) on L. vannamei juveniles reared for 21 days at low (5 g L^?1), medium (30 g L^?1) and high salinities (50 g L^?1). Juveniles grown at 5 g L^?1 had lower survival compared with controls (30 g L^?1) or shrimp grown at 50 g L^?1, but no significant effect on survival was observed as a result of HUFA enrichment. In contrast, growth was significantly lower for shrimp grown at 50 g L^?1, but this effect was compensated by the HUFA‐enriched diet. Osmotic pressure in haemolymph was affected by salinity, but not by HUFA enrichment. Shrimp fed HUFA‐enriched diets had significantly higher levels of eicosapentaenoic acid and docosahexaenoic acid in hepatopancreas and gills. These results demonstrate that growth at high salinities is enhanced with diets containing high HUFA levels, but that HUFA‐enriched diets have no effect on shrimp reared at low salinities.