First isolation of a novel thermostable antifungal peptide secreted by Aspergillus clavatus

A novel antifungal peptide produced by an indigenous fungal strain (VR) of Aspergillus clavatus was purified. The antifungal peptide was enriched in the supernatant after heat treatment at 70 degrees C. The thermostable character was exploited in the first purification step, as purified peptide was obtained after ultrafiltration and reverse phase-HPLC on C18 column application. The purified peptide named "AcAFP" for A. clavatus antifungal peptide, has molecular mass of 5773Da determined by MALDI-ToF spectrometry. The N-terminal sequence showed a notable identity to the limited family of antifungal peptides produced by ascomycetes fungi. The AcAFP activity remains intact even after heat treatment at 100 degrees C for 1h confirming its thermostability. It exhibits a strong inhibitory activity against mycelial growth of several serious human and plant pathogenic fungi: Fusariuym oxysporum, Fusarium solani, Aspergillus niger, Botrytis cinerea, Alternaria solani, whereas AcAFP did not affect yeast and bacterial growth.     

Production of Hexenol in a Two-Enzyme System: Kinetic Study and Modelling

In a two-enzyme system, successive action of hydroperoxide lyase from mint and yeast alcohol-dehydrogenase catalyses the conversion of hydroperoxy linolenic acid to hexenol. Kinetic behaviour was investigated separately for each enzyme: a lumped model based on the Michaelis-Menten approach shows the fate of the reactants in the system.     

Intérêt de l’étude de l’oxydation de l’ADN des spermatozoïdes par marquage de la 8-oxo-guanine en cytométrie en flux chez l’homme infertile

Oxidative stress in semen is essentially due to excessive production of oxygen-reactive species (ORS) essentially derived from leukocytes. DNA oxidation is due to the direct action of ORS which produce several adducts the most extensively studied of which is 8-oxo-guanine. Integrity of DNA is essential for the fertility of sperm and is an important subject of research for scientists and clinicians all over the world. Although evaluation of the global integrity of sperm DNA has considerably developed over recent years, few tests are available to document oxidative DNA damage. This study was designed to review the various tests of sperm DNA integrity commonly used in the literature and to present the results of our study on DNA oxidation with 8-oxo-guanine labelling by flow cytometry in infertile men. This study was based on 15 semen samples that were submitted to sperm analysis according to WHO guidelines, with determination of the leukocyte concentration by a cytochemical method revealing peroxidase in cytoplasmic granulations. The DNA oxidation study was performed with 8-oxo-guanine labelling by flow cytometry. Linear regression analysis showed a strong correlation between DNA oxidation and leukocyte count in the semen (p = 0.006, r = 0.7). A leukocyte cut-off of 250,000/ml of semen was associated with a significant increase of DNA oxidation (p = 0.03). 8-oxo-guanine can therefore be considered to be a biological marker of the direct action of oxidative stress on sperm DNA which appears to be susceptible to relatively low levels of ORS produced by leukocytes present at concentrations well below the limit of leukospermia defined by WHO.     

Gene–environment interactions between <Emphasis Type="Italic">ERCC2</Emphasis>, <Emphasis Type="Italic">ERCC3</Emphasis>, <Emphasis Type="Italic">XRCC1</Emphasis> and cadmium exposure in nasal polyposis disease

Gene–environment interactions have long been known to play an important role in complex disease aetiology, such as nasal polyposis (NP). The present study supports the concept that DNA repair gene polymorphisms play critical roles in modifying individual susceptibility to environmental diseases. In fact, we investigated the role of polymorphisms in DNA repair genes and cadmium as risk factors for Tunisian patients with NP. To the best of our knowledge, this is the first report on the impact of combined effects of cadmium and ERCC3 7122 A>G (rs4150407), ERCC2 Lys751Gln (rs13181) and XRCC1 Arg399Gln (rs25487) genes in the susceptibility to NP disease. Significant associations between the risk of developing NP disease and ERCC2 [odds ratio (OR)?=?2.0, 95 % confidence interval (CI)?=?1.1–3.7, p?=?0.023] and ERCC3 (OR?=?2.2, 95 % CI?=?1.2–4.1, p?=?0.013) genotypes polymorphisms were observed. Blood concentrations of Cd in NP patients (2.2 μg/L) were significantly higher than those of controls (0.5 μg/L). A significant interaction between ERCC3 (7122 A>G) polymorphism and blood-Cd levels (for the median of blood-Cd levels: OR?=?3.8, 95 % CI?=?1.3–10.8, p?=?0.014 and for the 75th percentiles of blood-Cd levels: OR?=?2.7, 95 % CI?=?1.1–7.2, p?=?0.041) was found in association with the risk of NP disease. In addition, when we stratified ERCC2, ERCC3 and XRCC1 polymorphism genotypes by the median and 75th percentiles of blood-Cd levels, we found also significant interactions between ERCC2 (Lys751Gln) and ERCC3 (7122 A>G) genotypes polymorphism and this metal in association with NP disease. However, no interaction was found between XRCC1 (Arg399Gln) polymorphism genotypes and Cd in association with NP disease.     

Catalytic properties of the immobilized Talaromyces thermophilus β-xylosidase and its use for xylose and xylooligosaccharides production

The present study explores the efficiency of Talaromyces thermophilus β-xylosidase, in the production of xylose and xylooligosaccharides. The β-xylosidase was immobilized by different methods namely ionic binding, entrapment and covalent coupling and using various carriers. Chitosan, pre-treated with glutaraldehyde, was selected as the best support material for β-xylosidase immobilization; it gave the highest immobilization and activity yields (94%, 87%, respectively) of initial activity, and also provided the highest stability, retaining 94% of its initial activity even after being recycled 25 times. Shifts in the optimal temperature and pH were observed for the immobilized β-xylosidase when compared to the free enzyme. The maximal activity obtained for the immobilized enzyme was achieved at pH 8.0 and 53 °C, whereas that for the free enzyme was obtained at pH 7.0 and 50 °C. The immobilized enzyme was more thermostable than the free β-xylosidase. We observed an increase of the K_m values of the free enzyme from 2.37 to 3.42 mM at the immobilized state. Native and immobilized β-xylosidase were found to be stimulated by Ca²⁺, Mn²⁺ and Co²⁺ and to be inhibited by Zn²⁺, Cu²⁺, Hg²⁺, Fe²⁺, EDTA and SDS. Immobilized enzyme was found to catalyze the reverse hydrolysis reaction, forming xylooligosaccharides in the presence of a high concentration of xylose. In order to examine the synergistic action of xylanase and β-xylosidase of T. thermophilus, these two enzymes were co-immobilized on chitosan. A continuous hydrolysis of 3% Oat spelt xylan at 50 °C was performed and better hydrolysis yields and higher amount of xylose was obtained.     

A two-enzyme system for the transformation of unsaturated oils to 9(S)-hydroperoxy fatty acids

A two-enzyme system involving a lipase from a Pseudomonassp. and an extract of potato tubers containing lipoxygenase was used to convert triacylglycerols to 9-hydroperoxy fatty acids. Highest yields (up to 25%) were obtained with 1 to 20 g l^–1 trilinolein as substrate after 5 h under O₂ at pH 6 and 25 °C. The product structure was confirmed by ¹H NMR, MS and IR.     

Studies on the ecology of actinomycetes in an agricultural soil amended with organic residues: II. Assessment of enzymatic activities of <Emphasis Type="Italic">Actinomycetales</Emphasis> isolates

The main objective of this investigation was to study the enzymatic activities of Actinomycetales strains isolated from an agricultural soil amended with different amounts of municipal solid waste compost (MSWC) or farmyard manure (FM). For this purpose, the hydrolytic activities of carboxymethyl cellulase, xylanase, pectinase, amylase, chitinase and protease were tested for 75 isolates of Sterptomyces, Amycolatopsis and Nocardioides from different sources (unamended soil, amended soil with FM or MSWC, FM and MSWC) at temperature ranging between 30 and 50°C. It was shown that the highest rate of enzymes producer’s strains was registered at 30°C, and decreased gradually to annul at 50°C, with the exception of the MSWC strains origin. It was also shown that the percentage of strains producers of enzymes isolated from soil amended with MSWC appeared higher than the one registered for those isolated from control and amended with FM soils. Application of MSWC increases the number of enzymes producer-actinomycetes in the soil and then it improves its fertility.     

Chemical Composition,Antioxidant Potential and Enzymes Inhibitory Properties of Globe Artichoke By‐Products

In this study, chemical composition and in vitro biological activities of artichoke by‐products (leaves, floral stems and bracts) issued from two Tunisian varieties were evaluated. Analysis was performed by means of high‐performance liquid chromatography with diode array detection coupled to electrospray ionization mass spectrometric (LC/DAD/ESI‐MS). Total phenolic (TPC) and flavonoid (TFC) contents as well as the antioxidant activity conducted by three complementary methods, DPPH, ABTS and FRAP tests, were performed for each sample. Enzyme inhibitory effects against acetylcholinesterase, butyrylcholinesterase and α‐amylase were also studied. Results showed that TPC and TFC varied according to variety as well as the plant part. Bracts presented the highest TPC values (10–15 mg GAE/g DW), while leaves were distinguished by the highest TFC values (52–58 mg EQ/g DW). In vitro assays showed that Violet d'Hyères bracts and Blanc d'Oran leaves present the most antioxidant activities (30.040 and 20.428 mgET/gDW, respectively, by the DPPH method). Leaves demonstrated the highest acetylcholinesterase and butyrylcholinesterase inhibitory effects. Moreover, all organs displayed a noticeable inhibition towards α‐amylase. LC/DAD/MS analysis revealed that artichoke by‐products are a potential source of biopharmaceuticals such as luteolin derivatives from leaves and mono/dicaffeoylquinic acids in the other parts. This research demonstrates that globe artichoke by‐products, unexploited in our country, are a promising source of natural health promoting compounds with potential applications in the food and pharmaceutical industries.     

Cloning, Molecular Characterization, and mRNA Expression of the Thermostable Family 3 β-Glucosidase from the Rare Fungus Stachybotrys microspora

The filamentous fungus Stachybotrys microspora possess a rich β-glucosidase system composed of five β-glucosidases. Three of them were already purified to homogeneity and characterized. In order to isolate the β-glucosidase genes from S. microspora and study their regulation, a PCR strategy using consensus primers was used as a first step. This approach enabled the isolation of three different fragments of family 3 β-glucosidase gene. A representative genomic library was constructed and probed with one amplified fragment gene belonging to family 3 of β-glucosidase. After two rounds of hybridization, seven clones were obtained and the analysis of DNA plasmids leads to the isolation of one clone (CF3) with the largest insert of 7 kb. The regulatory region shows multiple TC-rich elements characteristic of constitutive promoter, explaining the expression of this gene under glucose condition, as shown by zymogram and RT-PCR analysis. The tertiary structure of the deduced amino acid sequence of Smbgl3 was predicted and has shown three conserved domains: an (α/β)₈ triose phosphate isomerase (TIM) barrel, (α/β)₅ sandwich, and fibronectin type III domain involved in protein thermostability. Zymogram analysis highlighted such thermostable character of this novel β-glucosidase.