In this work, two biosurfactant-producing strains, Bacillus subtilis and Bacillus licheniformis, have been characterized. Both strains were able to grow at high salinity conditions and produce biosurfactants up to 10% NaCl. Both extracted-enriched biosurfactants showed good surface tension reduction of water, from 72 to 26-30 mN/m, low critical micelle concentration, and high resistance to pH and salinity. The potential of the two lipopeptide biosurfactants at inhibiting biofilm adhesion of pathogenic bacteria was demonstrated by using the MBEC device. The two biosurfactants showed interesting specific anti-adhesion activity being able to inhibit selectively biofilm formation of two pathogenic strains. In particular, Escherichia coli CFT073 and Staphylococcus aureus ATCC 29213 biofilm formation was decreased of 97% and 90%, respectively. The V9T14 biosurfactant active on the Gram-negative strain was ineffective against the Gram-positive and the opposite for the V19T21. This activity was observed either by coating the polystyrene surface or by adding the biosurfactant to the inoculum. Two fractions from each purified biosurfactant, obtained by flash chromatography, fractions (I) and (II), showed that fraction (II), belonging to fengycin-like family, was responsible for the anti-adhesion activity against biofilm of both strains.

Lipopeptide biosurfactants produced by the Bacillus licheniformis V9T14 strain showed an interesting anti-adhesion activity against biofilm formation of human pathogenic bacterial strains. The chemical characterisation of the crude extract of V9T14 strain was first developed through electrospray ionisation mass spectrometry (ESI-MS) and ESI-MS/MS direct infusions: two sets of molecular ion species belonging to the fengycin and surfactin families were revealed and their structures defined, interpreting their product ion spectra. The LC/ESI-MS analysis of the crude extract allowed to separate in different chromatogram ranges the homologues and the isoforms of the two lipopeptide families. The extract was then fractionated by silica gel chromatography in two main fractions, I and II. The purified biosurfactants were analysed through a new, rapid and suitable LC/ESI-MS/MS method, which allowed characterising the composition and the structures of the produced lipopeptides. LC/ESI-MS/MS analysis of fraction I showed the presence of C(13), C(14) and C(15) surfactin homologues, whose structures were confirmed by the product ion spectra of the sodiated molecules [M + Na](+) at m/z 1030, 1044 and 1058. LC/ESI-MS/MS analysis of fraction II confirmed the presence of two main fengycin isoforms, with the protonated molecules [M + H](+) at m/z 1478 and 1506 corresponding to C(17) fengycin A and C(17) fengycin B, respectively. Other homologues (C(14) to C(16)) were revealed and confirmed as belonging to fengycin A or B according to the retention times and the product ions generated, although with the same nominal mass. Finally, a relative percentage content of each homologue for both lipopeptides families in the whole extract was proposed.

No abstract

DmsD is a chaperone of the redox enzyme maturation protein family specifically required for biogenesis of DMSO reductase in Escherichia coli. It exists in multiple folding forms, all of which are capable of binding its known substrate, the twin-arginine leader sequence of the DmsA catalytic subunit. It is important for maturation of the reductase and targeting to the cytoplasmic membrane for translocation. Here, we demonstrate that DmsD exhibits an irreversible photobleaching phenomenon upon 280 nm excitation irradiation. The phenomenon is due to quenching of the tryptophan residues in DmsD and is dependent on its folding and conformation. We also show that a tryptophan residue involved in DmsA signal peptide binding (W87) is important for photobleaching of DmsD. Mutation of W87, or binding of the DmsA twin-arginine signal peptide to DmsD in the pocket that includes W72, W80, and W91 significantly affects the degree of photobleaching. This study highlights the advantage of a photobleaching phenomenon to study protein folding and conformation changes within a protein that was once considered unusable in fluorescence spectroscopy.

Nowadays, notwithstanding their nutritional and technological properties, food bioactive peptides from plant sources garner increasing attention for their ability to impart more than one beneficial effect on human health. Legumes, which stand out thanks to their high protein content, represent valuable sources of bioactive peptides. In this context, this study focused on the characterization of the potential pleotropic activity of two commercially available soybean (SH) and pea (PH) protein hydrolysates, respectively. Since the biological activity of a specific protein hydrolysate is strictly correlated with its chemical composition, the first aim of the study was to identify the compositions of the SH and PH peptides. Peptidomic analysis revealed that most of the identified peptides within both mixtures belong to storage proteins. Interestingly, according to the BIOPEP-UWM database, all the peptides contain more than one active motive with known inhibitory angiotensin converting enzyme (ACE) and dipeptidyl-dipeptidases (DPP)-IV sequences. Indeed, the results indicated that both SH and PH inhibit DPP-IV and ACE activity with a dose-response trend and IC50 values equal to 1.15 ± 0.004 and 1.33 ± 0.004 mg/mL, and 0.33 ± 0.01 and 0.61 ± 0.05 mg/mL, respectively. In addition, both hydrolysates reduced the activity of DPP-IV and ACE enzymes which are expressed on the surface of human intestinal Caco-2 cells. These findings clearly support that notion that SH and PH may represent new ingredients with anti-diabetic and hypotensive effects for the development of innovative multifunctional foods and/or nutraceuticals for the prevention of metabolic syndrome.

Biological contamination of surfaces, both in industry and in health care, plays an important role as a potential vector of disease transmission. Metals have been described to be effective antibiofilm agents, and the efficacy of silver ions as a disinfectant has been known for centuries. The activity of AgNO3 combined with the lipopeptide biosurfactant V9T14 has been studied against a preformed Escherichia coli biofilm on the Calgary Biofilm Device. Results indicated that the activity of silver can be synergistically enhanced by the presence of V9T14, both allowing for a reduction in the quantity of silver used and for greater antimicrobial activity. The concentration of silver needed to obtain this reduction in the silver-biosurfactant solution was from 129- to 258-fold less than the concentration of silver alone. To our knowledge, this is the first time that a synergistic interaction between a lipopeptide biosurfactant and silver has been observed.

The objective of the study was the evaluation of the potential pleiotropic effect of a commercial casein hydrolysate (CH). After an analysis of the composition, the BIOPEP-UWM database suggested that these peptides contained numerous sequences with potential inhibitory activities on angiotensin converting enzyme (ACE) and dipeptidyl-peptidase IV (DPP-IV). The anti-diabetic and anti-hypertensive effects of these peptides were thus assessed using either cell-free or cell-based assays. In the cell-free system, CH displayed inhibitory properties against DPP-IV (IC50 value equal to 0.38 ± 0.01 mg/mL) and ACE (IC50 value equal to 0.39 ± 0.01 mg/mL). Further, CH reduced the DPP-IV and ACE activities expressed by human intestinal Caco-2 cells by 61.10 ± 1.70% and 76.90 ± 4.47%, respectively, versus untreated cells, after 6 h of treatment at the concentration of 5 mg/mL. This first demonstration of the multifunctional behavior of this material suggests that it may become an anti-diabetic and/or anti-hypertensive ingredient to be included in the formulation of different functional food or nutraceutics.

Hempseed (Cannabis sativa) is one of the most promising sources of plant proteins. It contains approximately 24% (w/w) protein, and edestin accounts for approximately 60–80% (w/w) of its total proteins. In a framework of research aimed at fostering the proteins recovered from the press cake by-products generated after the extraction of hempseed oil, two hempseed protein hydrolysates (HH1 and HH2) were produced at an industrial level using a mixture of different enzymes from Aspergillus niger, Aspergillus oryzae, and Bacillus licheniformis for different times (5 h and 18 h). Using a combination of different direct antioxidant tests (DPPH, TEAC, FRAP, and ORAC assays, respectively), it has been demonstrated that HHs exert potent, direct antioxidant activity. A crucial feature of bioactive peptides is their intestinal bioavailability; for this reason, in order to solve this peculiar issue, the ability of HH peptides to be transported by differentiated human intestinal Caco-2 cells has been evaluated. Notably, by using mass spectrometry analysis (HPLC Chip ESI-MS/MS), the stable peptides transported by intestinal cells have been identified, and dedicated experiments confirmed that the trans-epithelial transported HH peptide mixtures retain their antioxidant activity, suggesting that these hempseed hydrolysates may be considered sustainable antioxidant ingredients to be exploited for further application, i.e., nutraceutical and/or food industries.