Complete1H NMR assignments of synthetic glycopeptides from the carbohydrate-protein linkage region of serglycins

We present complete¹H NMR assignments for two synthetic glycopeptides representative of the carbohydrateprotein linkage region of serglycin proteoglycans. The peptides are: Ser(Galp-Xylp)-Gly-Ser-Gly-Ser(Galp-Xylp)-Gly and, Ser(Galp-Xylp)-Gly-Ser(Galp-Xylp)-Gly-Ser(Galp-Xylp)-Gly. A number of 2D NMR spectra together with a 3D NOESY-TOCSY spectrum were acquired at 600 MHz to complete the assignments of the glycopeptides dissolved in water with 40% trifluoroethanol. Preliminary analysis of the NMR data suggests folded structures for the glycopeptides.A preliminary account of this work was presented at an International Symposium held at the University of Alabama at Birmingham in November, 1994 on the occasion of the 65th birthday of Professor Lennart Rodén.     

Evidence of an insulin generated pyruvate dehydrogenase stimulating factor in rat brain plasma membranes

1. The results of this study indicates that the binding of insulin to brain plasma membranes activates a membrane protease which, by a trypsin like mechanism, produces a soluble factor that modulates the PDH behaviour when added to brain mitochondria. 2. The supernatant from brain plasma membranes incubated with 0.5 mg/ml trypsin added to mitochondria increases PDH activity levels and cancels PDH inhibition by NaF, as has already been seen when the plasma membranes are incubated with 25 microU/ml insulin. No such effects are obtained when the incubation is run out with 0.5 mg/ml chymotrypsin. 3. The supernatants from insulin or trypsin treated plasma membranes retain their activating properties on mitochondrial PDH also after dansylation; from these preparations a dansylated active on PDH material was separated by monodimensional chromatography on HPTLC silica Gel plates, using chloroform/1-butanol (93:7 v/v) as a solvent. 4. Insulin incubation of plasma membranes pretreated with protease inhibitors (leupeptin, phenylmethylsulfonylfluoride) or with exogenous trypsin, but not chymotrypsin substrates (esters of arginine and tyrosine) yields an inactive supernatant on PDH. 5. Insulin treated plasma membrane supernatants lose all stimulating properties on PDH after incubation for 1 hr with 2 mg/ml trypsin or chymotrypsin.     

Increased kynurenine-to-tryptophan ratio in the serum of patients infected with SARS-CoV2: An observational cohort study.

Immune dysregulation is a hallmark of patients infected by SARS-CoV2 and the balance between immune reactivity and tolerance is a key determinant of all stages of infection, including the excessive inflammatory state causing the acute respiratory distress syndrome. The kynurenine pathway (KP) of tryptophan (Trp) metabolism is activated by pro-inflammatory cytokines and drives mechanisms of immune tolerance. We examined the state of activation of the KP by measuring the Kyn:Trp ratio in the serum of healthy subjects (n = 239), and SARS-CoV2-negative (n = 305) and -positive patients (n = 89). Patients were recruited at the Emergency Room of St. Andrea Hospital (Rome, Italy). Kyn and Trp serum levels were assessed by HPLC/MS-MS. Compared to healthy controls, both SARS-CoV2-negative and -positive patients showed an increase in the Kyn:Trp ratio. The increase was larger in SARS-CoV2-positive patients, with a significant difference between SARS-CoV2-positive and -negative patients. In addition, the increase was more prominent in males, and positively correlated with age and severity of SARS-CoV2 infection, categorized as follows: 1 = no need for intensive care unit (ICU); 2 ≤ 3 weeks spent in ICU; 3 ≥ 3 weeks spent in ICU; and 4 = death. The highest Kyn:Trp values were found in SARS-CoV2-positive patients with severe lymphopenia. These findings suggest that the Kyn:Trp ratio reflects the level of inflammation associated with SARS-CoV2 infection, and, therefore, might represent a valuable biomarker for therapeutic intervention.     

Isoflavone synthase (IFS) gene phylogeny in Trifolium species associated with plant isoflavone contents

This study presents the results of the identification and quantification of 12 isoflavones (prunetin, irilone, pseudobaptigenin, glycitein, daidzin, genistin, daidzein, pratensein, puerarin, biochanin A, formononetin and genistein) in 23 species of Trifolium (T. arvense, T. pratense, T. ligusticum, T. striatum, T. lappaceum, T. angustifolium, T. hirtum, T. subterraneum, T. isthmocarpum, T. stellatum, T. mutabile, T. strictum, T. fragiferum, T. alexandrinum, T. tomentosum. T. nigrescens subsp. petrisavii, T. nigrescens, T. glomeratum, T. subterraneum subsp. brachycalycinum, T. cherleri, T. resupinatum, T. campestre and T. repens). Isoflavones were extracted by an MSPD method and analyzed with HPLC coupled with a diode-array detector. The evaluation of molecular phylogeny of the IFS gene and the relation with isoflavone content was also performed. Five species (T. subterraneum subsp. brachycalycinum, T. alexandrinum, T. pratense, T. subterraneum and T. lappaceum) were identified with high levels of biochanin A (431–83 mg/kg), formononetin (72–365 mg/kg) and genistein (9–509 mg/kg), which could be utilized as alternative sources for the nutraceutical industry. Genetic phylogeny for the IFS gene was found in the species studied, with 20 out of 23 species having been divided into two clades, while the remaining three were genetically distant. Based on our results, we confirm the direct correlation between IFS gene polymorphism and isoflavones content in species of Trifolium particularly noted for formononetin. Therefore, the IFS gene can be utilized for screening Trifolium genotypes for formononetin. The relation of the three isoflavones' contents and the molecular phylogeny of plants determined by the IFS sequences, as a screening marker for plants with high isoflavone contents in Trifolium species, are to the best of our knowledge described for the first time.     

Application of a SSR‐GBS marker system on investigation of European Hedgehog species and their hybrid zone dynamics

By applying second‐generation sequencing technologies to microsatellite genotyping, sequence information is produced which can result in high‐resolution population genetics analysis populations and increased replicability between runs and laboratories. In the present study, we establish an approach to study the genetic structure patterns of two European hedgehog species Erinaceaus europaeus and E. roumanicus. These species are usually associated with human settlements and are good models to study anthropogenic impacts on the genetic diversity of wild populations. The short sequence repeats genotyping by sequence (SSR‐GBS) method presented uses amplicon sequences to determine genotypes for which allelic variants can be defined according to both length and single nucleotide polymorphisms (SNPs). To evaluate whether complete sequence information improved genetic structure definition, we compared this information with datasets based solely on length information. We identified a total of 42 markers which were successfully amplified in both species. Overall, genotyping based on complete sequence information resulted in a higher number of alleles, as well as greater genetic diversity and differentiation between species. Additionally, the structure patterns were slightly clearer with a division between both species and some potential hybrids. There was some degree of genetic structure within species, although only in E. roumanicus was this related to geographical distance. The statistically significant results obtained by SSR‐GBS demonstrate that it is superior to electrophoresis‐based methods for SSR genotyping. Moreover, the greater reproducibility and throughput with lower effort which can be obtained with SSR‐GBS and the possibility to include degraded DNA into the analysis, allow for continued relevance of SSR markers during the genomic era.     

Ezrin is essential for epithelial organization and villus morphogenesis in the developing intestine

Ezrin, Radixin, and Moesin (the ERM proteins) supply regulated linkage between membrane proteins and the actin cytoskeleton. The study of mammalian ERM proteins has been hampered by presumed functional overlap. We have found that Ezrin, the only ERM detected in epithelial cells of the developing intestine, provides an essential role in configuring the mouse intestinal epithelium. Surprisingly, Ezrin is not absolutely required for the formation of brush border microvilli or for the establishment or maintenance of epithelial polarity. Instead, Ezrin organizes the apical terminal web region, which is critical for the poorly understood process of de novo lumen formation and expansion during villus morphogenesis. Our data also suggest that Ezrin controls the localization and/or function of certain apical membrane proteins that support normal intestinal function. These in vivo studies highlight the critical function of Ezrin in the formation of a multicellular epithelium rather than an individual epithelial cell.     

Yeast production from virgin grape marc

An alternative utilization of virgin grape marc (VGM) to produce SCP from S. ceretisiae is reported. A simple extraction method of fresh grape marc produces a sugar-rich solution: through fed-batch fermentation, a high-value yeast biomass instead of a low-value product like ethanol can be produced. Productivity and quality of yeast are similar to these obtainable from molasses. The convenience of yeast production from VGM is briefly discussed; it appears of great interest in south Italy and generally in grape-producing countries, specially if these lack relevant sources of fermentable sugars.