The crystal structure of Rv0813c from Mycobacterium tuberculosis reveals a new family of fatty acid-binding protein-like proteins in bacteria

The gene Rv0813c from Mycobacterium tuberculosis, which codes for a hypothetical protein of unknown function, is conserved within the order Actinomycetales but absent elsewhere. The crystal structure of Rv0813c reveals a new family of proteins that resemble the fatty acid-binding proteins (FABPs) found in eukaryotes. Rv0813c adopts the 10-stranded beta-barrel fold typical of FABPs but lacks the double-helix insert that covers the entry to the binding site in the eukaryotic proteins. The barrel encloses a deep cavity, at the bottom of which a small cyclic ligand was found to bind to the hydroxyl group of Tyr192. This residue is part of a conserved Arg-X-Tyr motif much like the triad that binds the carboxylate group of fatty acids in FABPs. Most of the residues forming the internal surface of the cavity are conserved in homologous protein sequences found in CG-rich prokaryotes, strongly suggesting that Rv0813c is a member of a new family of bacterial FABP-like proteins that may have roles in the recognition, transport, and/or storage of small molecules in the bacterial cytosol.     

Cuticular waxes from ivy leaves (Hedera helix L.): analysis of high-molecular-weight esters

Soluble waxes were extracted from the cuticle of ivy (Hedera helix L.) leaves with dichloromethane in a yield of ca. 13%. The cuticular waxes were directly analysed by GC-MS, high-temperature GC-MS and ESI-MS/MS. The GC-MS analysis showed mostly n-alkanols (45.3%), monoacids (18.8%), triterpenes (9.7%), n-aldehydes (8.7%) and n-alkanes (7.7%). The high-temperature GC-MS and the ESI-MS/MS analyses showed the presence of ester waxes, namely alkyl alkanoates and alkyl coumarates. Alkyl alkanoates comprised esters of the hexadecanoic acid with n-alkanols ranging from C16 to C34. Alkyl coumarates included esters of coumaric acid with n-alkanols ranging from C16 to C32. The cuticular waxes were hydrolysed and the resulting organic and aqueous phases analysed by GC-MS. The hydrolysate showed a major increase in the quantities of n-alkanols, hexadecanoic acid and coumaric acid derived from the alkyl and acyl moieties from the ester waxes. A content of ester waxes of 38% was estimated based on the results from the GC-MS analysis of the non-hydrolysed and hydrolysed cuticular waxes. Alkyl alkanoates were analysed by ESI-MS/MS as [M + Li]+ adduct ions and the alkyl coumarates as [M - H]- deprotonated ions. The ESI-MS/MS analysis allowed the detection of a wider range of ester waxes than high-temperature GC-MS, and was shown to be a useful technique for the qualitative analysis of ester waxes from plant cuticles.     

Utilization of monocyclic aromatic hydrocarbons individually and in mixture by bacteria isolated from petroleum-contaminated soil

The fate of benzene, ethylbenzene, toluene, xylenes (BTEX) compounds through biodegradation was investigated using two different bacteria, Ralstonia picketti (BP-20) and Alcaligenes piechaudii (CZOR L-1B). These bacteria were isolated from extremely polluted soils contaminated with petroleum hydrocarbons. PCR and Fatty Acid Methyl Ester (FAME) were used to identify the isolates. In this study, BTEX biodegradation, applied as a mixture or as individual compounds by the bacteria was evaluated. Both bacteria were shown to degrade each of the BTEX compounds individually and in mixture. However, Alcaligenes piechaudii was a better degrader of BTEXs both in the mixture and individually. Differences between BTEX biodegradation in the mixture and individually were observed, especially in the case of benzene. The degradation of all BTEXs in the mixture was lower than the degradation of individual compounds for both bacteria tested. In the all experiments, toluene and m + p- xylenes were better removed than the other BTEXs. No intermediates of biodegradation were detected. Biosurfactant production was observed by culture techniques. In addition, 3-hydroxy fatty acids, important in biosurfactant production, were observed by FAME analysis. The test results indicate that the bacteria could contribute to bioremediation of aromatic hydrocarbon pollution.     

Maternal Lead Exposure Produces Long-Term Enhancement of Dopaminergic Reactivity in Rat Offspring

To determine the effect of prenatal lead exposure on brain monoaminergic systems, pregnant rats were given tap water containing 250 ppm lead acetate, for the duration of pregnancy, while tap water without lead (Pb²⁺) was substituted at birth. Control rats were derived from dams that consumed tap water during pregnancy, and had no exposure to lead afterwards. At 12 weeks after birth, Pb²⁺ content of brain cortex was increased 3- to 4-fold (P < 0.05). At this time the endogenous striatal levels of 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid were 19% lower in Pb²⁺ exposed rats (P < 0.05), while there was no change in the striatal level of dopamine (DA), noradrenaline, 3,4-dihydroxyphenylglycol, serotonin (5-HT) and 5-hydroxyindoleacetic acid (HPLC/ED). Also there was no change in these monoamines and metabolites in the prefrontal cortex of Pb²⁺ exposed rats. However, turnover of 5-HT in prefrontal cortex, as indicated by 5-hydroxytryptophan accumulation 30 min after acute treatment with the decarboxylase inhibitor NSD-1015 (100 mg/kg IP), was lower in the Pb²⁺ exposed rats. In the striatum AMPH-induced (1 mg/kg IP) turnover of DA, evidenced as L-DOPA accumulation after NSD-1015, was increased to a lesser extent in the Pb²⁺ exposed rats (P < 0.05). The nitric oxide synthase inhibitor 7-nitroindazole (10 mg/kg IP) attenuated the latter effect, indicating that neuronal NO mediates this AMPH effect, at least in part. Moreover, DA D₂ receptor sensitivity developed in Pb²⁺ exposed rats, as evidenced by enhanced quinpirole-induced yawning activity and enhanced quinpirole-induced locomotor activity (each, P < 0.05). These findings indicate that ontogenetic exposure to lead can have consequences on monoaminergic neuronal function at an adult stage of life, generally promoting accentuated behavioral effects of direct and indirect monoaminergic agonists, and related to increased dopamine turnover in basal ganglia. Special issue dedicated to Dr. Moussa Youdim.     

Rab27b regulates mast cell granule dynamics and secretion

The Rab GTPase family regulates membrane domain organization and vesicular transport pathways. Recent studies indicate that one member of the family, Rab27a, regulates transport of lysosome-related organelles in specialized cells, such as melanosomes and lytic granules. Very little is known about the related isoform, Rab27b. Here we used genetically modified mice to study the involvement of the Rab27 proteins in mast cells, which play key roles in allergic responses. Both Rab27a and Rab27b isoforms are expressed in bone marrow-derived mast cells (BMMC) and localize to secretory granules. Nevertheless, secretory defects as measured by beta-hexosaminidase release in vitro and passive cutaneous anaphylaxis in vivo were found only in Rab27b and double Rab27 knockout (KO) mice. Immunofluorescence studies suggest that a subset of Rab27b and double Rab27-deficient BMMCs exhibit mild clustering of granules. Quantitative analysis of live-cell time-lapse imaging revealed that BMMCs derived from double Rab27 KO mice showed almost 10-fold increase in granules exhibiting fast movement (>1.5 microm/s), which could be disrupted by nocodazole. These results suggest that Rab27 proteins, particularly Rab27b, play a crucial role in mast cell degranulation and that their action regulates the transition from microtubule to actin-based motility.     

Corynebacterium diphtheriae putative tellurite-resistance protein (CDCE8392_0813) contributes to the intracellular survival in human epithelial cells and lethality of Caenorhabditis elegans

Corynebacterium diphtheriae, the aetiologic agent of diphtheria, also represents a global medical challenge because of the existence of invasive strains as causative agents of systemic infections. Although tellurite (TeO3^2-) is toxic to most microorganisms, TeO3^2--resistant bacteria, including C. diphtheriae, exist in nature. The presence of TeO3^2--resistance (Te^R) determinants in pathogenic bacteria might provide selective advantages in the natural environment. In the present study, we investigated the role of the putative Te^R determinant (CDCE8392_813gene) in the virulence attributes of diphtheria bacilli. The disruption of CDCE8392_0813 gene expression in the LDCIC-L1 mutant increased susceptibility to TeO3^2- and reactive oxygen species (hydrogen peroxide), but not to other antimicrobial agents. The LDCIC-L1 mutant also showed a decrease in both the lethality of Caenorhabditis elegans and the survival inside of human epithelial cells compared to wild-type strain. Conversely, the haemagglutinating activity and adherence to and formation of biofilms on different abiotic surfaces were not regulated through the CDCE8392_0813 gene. In conclusion, the CDCE8392_813 gene contributes to the Te^R and pathogenic potential of C. diphtheriae.     

Reliability-based life cycle assessment for future solid waste management alternatives in Portugal

Background, aim, and scope  

This paper presents a study related to the application of the reliability-based life cycle assessment (LCA) to assess different alternatives for solid waste management in the Setúbal peninsula, Portugal. The current system includes waste collection, transport, sorting, recycling, and mechanical and biological treatment (MBT) by means of aerobic treatment and landfill. In addition, some future expansion plans are discussed.     

Evaluation of hexose and pentose in pre-cultivation of <Emphasis Type="Italic">Candida guilliermondii</Emphasis> on the key enzymes for xylitol production in sugarcane hemicellulosic hydrolysate

The evaluation of hexose and pentose in pre-cultivation of Candida guilliermondii FTI 20037 yeast on xylose reductase (XR) and xylitol dehydrogenase (XDH) enzymes activities was performed during fermentation in sugarcane bagasse hemicellulosic hydrolysate. The xylitol production was evaluated by using cells previously growth in 30.0 gl^?1 xylose, 30.0 gl^?1 glucose and in both sugars mixture (30.0 gl^?1 xylose and 2.0 gl^?1 glucose). The vacuum evaporated hydrolysate (80 gl^?1) was detoxificated by ion exchange resin (A-860S; A500PS and C-150-Purolite^®). The total phenolic compounds and acetic acid were 93.0 and 64.9%, respectively, removed by the resin hydrolysate treatment. All experiments were carried out in Erlenmeyer flasks at 200 rpm, 30°C. The maximum XR (0.618 Umg _Prot ^?1 ) and XDH (0.783 Umg _Prot ^?1 ) enzymes activities was obtained using inoculum previously growth in both sugars mixture. The highest cell concentration (10.6 gl^?1) was obtained with inoculum pre-cultivated in the glucose. However, the xylitol yield and xylitol volumetric productivity were favored using the xylose as carbon source. In this case, it was observed maximum xylose (81%) and acetic acid (100%) consumption. It is very important to point out that maximum enzymatic activities were obtained when the mixture of sugars was used as carbon source of inoculum, while the highest fermentative parameters were obtained when xylose was used.     

Biogenesis of melanosomes - the chessboard of pigmentation

Melanosomes are lysosome-related organelles in retinal pigment epithelial cells and epidermal melanocytes in which melanin pigments are synthesized and stored. Melanosomes are generated by multistep processes in which an immature unpigmented organelle forms and then subsequently matures. Such maturation requires inter-organellar transport of protein cargos required for pigment synthesis but also recruitment of effector proteins necessary for the correct transport of melanosomes to the cell periphery. Several studies have started to unravel the main pathways and mechanisms exploited by melanosomal proteins involved in melanosome structure and melanin synthesis. A major unexpected finding seen early in melanosome biogenesis showed the similarities between the fibrillar sheets of premelanosomes and amyloid fibrils. Late steps of melanosome formation are dependent on pathways regulated by proteins encoded by genes mutated in genetic diseases such as the Hermansky-Pudlak Syndrom (HPS) and different types of albinism. Altogether the findings from the past recent years have started to unravel how specialized cells integrate unique and ubiquitous molecular mechanisms in subverting the endosomal system to generate cell-type specific structures and their associated functions. Further dissection of the melanosomal system will likely shed light not only on the biogenesis of lysosome-related organelles but also on general aspects of vesicular transport in the endosomal system.