Peptide AS-48: prototype of a new class of cyclic bacteriocins

After the discovery of bacteriocin AS-48, a 70-residue cyclic peptide produced by Enterococcus faecalis subsp. liquefaciens, some naturally-occurring cyclic proteins from bacteria have been reported. AS-48 is encoded by the 68-kb pheromone-responsive plasmid pMB2, and the gene cluster involved in production and immunity has been identified and sequenced. This peptide exerts a bactericidal action on sensitive cells (most of the Gram-positive and some Gram-negative bacteria). Its target is the cytoplasmic membrane, in which it opens pores, leading to the dissipation of the proton motive force and cell death, a mechanism similar to that proposed for the action of defensins or, most generally, cationic antibacterial peptides. This fact, together with its remarkable stability and solubility over a wide pH range, suggest that this bacteriocin could be a good candidate as a natural food preservative. The amino acid composition of purified AS-48 shows the absence of modified or dehydrated residues, making it clearly different from lantibiotics. Bacteriocin AS-48 also differs from defensins in that it does not contain cysteines and consequently no disulfide bridges, which makes is high stability even more remarkable. Composition analysis of AS-48 shows a high proportion of basic to acidic amino acids, conferring to this peptide a strong basic character, with an isoelectric point close to 10.5. Determination of the AS-48 structural gene DNA sequence, together with the sequences of AS-48 protease digestion fragments and mass spectrometry determinations, allowed us to determine unambiguously the cyclic structure of the molecule, being the first example of a posttranslational modification in which a cyclic structure arises from a "head-to-tail" linkage. We have solved the three-dimensional structure of AS-48 in solution, and it consists of a globular arrangement of five alpha-helices enclosing a compact hydrophobic core. Interestingly, the head-to-tail peptide link between Trp-70 and Met-1 lies in the middle of alpha-helix 5, which is shown to have a pronounced effect on the stability of the three-dimensional structure. Analysis of structure-function relationship allowed us to propose models to understand the aspects of the molecular function of AS-48. The purpose of this work is to review recent developments in our understanding about the biochemical and biological characteristics and structure of this unusual type of bacteriocin.     

A simple approach to sensor discovery and fabrication on self-assembled monolayers on glass

Self-assembled monolayers (SAMs) on glass were used as a platform to sequentially deposit fluorophores and small molecules for ion sensing. The preorganization provided by the surface avoids the need for complex receptor design, allowing for a combinatorial approach to sensing systems based on small molecules. The resulting libraries are easily measured and show varied responses to a series of both cations and anions. This technology is transferable from the macro- to the microscale both via microcontact printing (microCP), where the fluorophore is printed onto a glass surface, and via direct attachment of the fluorophore to microchannel walls. The ease of miniaturization of this technology may make the generation of a wide variety of simple yet efficient microarrays possible.     

Solid-phase microextraction and gas chromatography-mass spectrometry of volatile compounds from avocado puree after microwave processing

Microwave processing offers an alternative to blanch fruits and vegetables, since the application of high temperature and short time often results in minimum damage. An experimental design was used to investigate the effect of microwave time, pH, and avocado leaves (independent variables) on avocado flavor (response) using solid-phase microextraction (SPME)-GC-MS. Among the fully characterized flavor volatiles, 19 compounds were derived from lipid oxidation and only 4 from the avocado leaves. The main components derived from lipids were aldehydes, ketones and alcohols. Terpenoids, estragole, and 2-hexenal [E] were volatiles derived from avocado leaves. When leaves were added to fresh and microwaved avocado terpenoids and 2-hexenal [E]/hexanal ratio increased, this behavior was considered to have a positive effect on the sensorial quality of the product. From the statistical analysis of the experimental design, it was possible to determinate that the most important factors influencing the abundance of flavor compounds derived from lipids were microwave time and pH. Maximum values of these compounds were detected at high levels of microwave time and low values of pH. On the other hand, response surface of terpenoids and estragole showed an increment when microwave time and avocado leaf was increased. The region of optimum response was 30 s microwave time, pH 5.5, and 1% of avocado leaf.     

Holographic phase-shift measurement during development of a fixed grating in lithium niobate crystals

We report what is believed to be the first direct measurement of the grating phase-shift evolution during white-light illumination for the development of a fixed grating in an Fe-doped lithium niobate crystal. Stabilized holographic recording is shown to be essential for such measurements. Experimental data are in good agreement with theory and allow computation of the relevant material parameters for the sample under analysis. The results are of the utmost relevance for understanding the advantageous behavior of oxidized samples in hologram fixing.     

Complexation of phenolic guests by endo- and exo-hydrogen-bonded receptors

This article describes the complexation of phenol derivatives by hydrogen-bonded receptors. These phenol receptors are formed by self-assembly of calix[4]arene dimelamine or tetramelamine derivatives with 5,5-diethylbarbiturate (DEB) or cyanurate derivatives (CYA). The double rosette assemblies 3(3).(DEB)6/(CYA)6 have their phenol-binding functionalities (ureido groups) at the top and at the bottom of the double rosette (exo-receptors). The tetrarosette assemblies 4(3).(DEB)12/(CYA)12 form a cavity with binding sites between the two double rosettes for guest encapsulation (endo-receptors). An intrinsic binding constant Ka of 202 M-1 and 286 M-1 for the binding of 4-nitrophenol to the ureido functionalized exo- and endo-receptors, respectively, was observed. For the exo-receptor a 1:6 stoichiometry was observed while for the endo-receptor 1:4 binding stoichiometry was determined by Job plot and MALDI-TOF MS. The important role that the hydroxy group's acidity plays in the complexation of 4-nitrophenol is clarified by binding studies with different phenol derivatives. The hydrogen-bonded receptors showed a much smaller response towards less acidic phenol derivatives.     

On-fiber photodegradation after solid-phase microextraction of p,p'-DDT and two of its major photoproducts,p,p'-DDE and p,p'-DDD

The potential of performing photochemical studies in solid phase microextraction (SPME) fibers, "photo-SPME", to study the photodegradation of p,p'-DDT and two of its major degradation products, p,p'-DDE and p,p'-DDD, is shown. Analyses were carried out by gas chromatography mass spectroscopy detection. DDT was extracted from aqueous solutions using five different commercial coatings. The fibers were then exposed to UV light emitted by a low-pressure mercury lamp. After 30 min of irradiation, the degradation of DDT only occurred in polydimethylsiloxane fibers. The on-fiber degradation kinetics of p,p'-DDT was studied from 2 to 60 min. A large number of photoproducts were generated and their kinetic behavior was studied. In order to clarify the possible photoreaction pathways for DDT, individual water solutions containing p,p'-DDD or p.p'-DDE were prepared and photo-SPME was performed for each compound at different irradiation times. On the basis of the photoproducts identified, some photodegradation pathways are proposed. Finally, aqueous photodegradation studies followed by SPME were performed and compared to the photo-SPME. This work will show the enormous potential of photo-SPME to perform photodegradation studies.     

Indirect automatic atomic absorption determination of sulphate based on its precipitation with barium

A continuous precipitation flame atomization atomic absorption spectrometric method for the indirect determination of sulphate is proposed. By using a reversed flow-injection configuration for precipitation as barium sulphate, this anion was determined in the range 10–150 g/ml, with a relative standard deviation of 2.6%. The sensitivity was increased by a factor of seven by using a nitrous oxide-acetylene flame to measure barium.     

Spectrophotometric determination of cadmium with uramyldiacetic acid

A procedure for the determination of cadmium(II) by uv spectrophotometry is proposed. This metal reacts at pH 6 with uramyldiacetic acid (UDA) to give a complex with a maximal absorbance at 265 nm and follows Beer's law in the range 0.12–3.6 μg Cd(II)/ml, the optimal interval being from 0.6 to 3.3 μg Cd(II)/ml. The sensitivity is 0.011 μg cm^?2. Through this study the stoichiometry of the chelate compound, interfering elements, and the reproducibility of the method were evaluated. The method has been applied to the determination of cadmium in biological samples and a white metal.     

Intracrystalline diffusion in metal organic framework during heterogeneous catalysis: influence of particle size on the activity of MIL-100 (Fe) for oxidation reactions

Three MIL-100 (Fe) samples differing in average crystal size (from 60-70 to >400 nm) have been synthesized by microwave heating using three HF/Fe(3+) ratios. Oxidation of diphenylmethane with tert-butylhydroperoxide (TBHP) and thiophenol with oxygen are catalyzed by three MIL-100 (Fe) samples with similar reaction rates regardless of its average particle size. In contrast, the activity of the three MIL-100 (Fe) samples for the oxidation of bulky triphenylmethane by TBHP largely depends on the average crystal size of the sample: the smaller the average particle size, the larger the initial reaction rate of triphenylmethane oxidation. These results show that diffusion limitation takes place on MOF catalysis depending on the substrate size and provides indirect evidence that these reactions take place inside the intracrystalline space of the porous catalysts.