The entire N-terminal half of TatC is involved in twin-arginine precursor binding

Translocation of twin-arginine precursor proteins across the cytoplasmic membrane of Escherichia coli requires the three membrane proteins TatA, TatB, and TatC. TatC and TatB were shown to be involved in precursor binding. We have analyzed in vitro a number of single alanine substitutions in tatC that were previously shown to compromise in vivo the function of the Tat translocase. All tatC mutants that were defective in precursor translocation into cytoplasmic membrane vesicles concomitantly interfered with precursor binding not only to TatC but also to TatB. Hence structural changes of TatC that affect precursor targeting simultaneously abolish engagement of the twin-arginine signal sequence with TatB and block the formation of a functional Tat translocase. Since these phenotypes were observed for tatC mutations spread over the first half of TatC, this entire part of the molecule must globally be involved in precursor binding.     

Effects of selected pharmaceutical products on phagocytic activity in Elliptio complanata mussels

Municipal wastewaters are recognized as a major source of pharmaceutical and personal care products to the aquatic environment, thereby exposing biota to unknown chronic effects. This study sought to examine the immunotoxic effects of pharmaceutical and urban waste products on the freshwater mussel Elliptio complanata. Hemolymph samples were collected and treated in vitro with increasing concentrations of various drugs (bezafibrate, carbamazepine, fluoxetine, gemfibrozil, morphine, naproxen, novobiocin, oxytetracycline, sulfamethazole, sulfapyridine and trimethoprim) and urban waste related chemicals (coprostanol, caffeine, cotinine) for 24 h at 15 degrees C. In a parallel experiment, mussels were caged and placed in two final aeration lagoons for the treatment of domestic wastewaters. At the end of the exposure period, hemolymphs were tested for phagocytic activity, intracellular esterase activity, cell adherence and lipid peroxidation (LPO). The products that most increased phagocytosis were bezafibrate, gemfibrozil and trimethoprim, while novobiocin and morphine reduced its activity. Intracellular esterase activity was reduced most strongly with sulfamethazole, novobiocin, gemfibrozil, bezafibrate and carbamazepine. Cell adherence was decreased by oxytetracycline, novobiocin and naproxen, and increased by gemfibrozil, bezafibrate and sulfapyridine. Exposure to these products also modulated LPO in hemocytes. Coprostanol and naproxen were more potent to reduce LPO while novobiocin and sulfapyridine were the most potent to induce LPO. The potential to induce LPO was positively correlated with the number of functional groups on the molecule (i.e., its nucleophilicity). Mussels exposed to domestic wastewater treatment plant aeration lagoons had decreased intracellular esterase and phagocytic activity as well, suggesting immunosuppression. PPCPs (pharmaceuticals and personal care products) that are recognized to disrupt cytokine signalling network by the nitric oxide pathway and cell permeability were generally the most potent ones. The data suggest that PPCPs have the potential to cause adverse effects on the immune system of bivalves.     

A Technique To Quantify the Population Size and Composition of the Biofilm Component in Communities of Bacteria in the Phyllosphere

The presence of microbial biofilms in the phyllosphere of terrestrial plants has recently been demonstrated, but few techniques to study biofilms associated with living plant tissues are available. Here we report a technique to estimate the proportion of the bacterial population on leaves that is assembled in biofilms and to quantitatively isolate bacteria from the biofilm and nonbiofilm (solitary) components of phyllosphere microbial communities. This technique is based on removal of bacteria from leaves by gentle washing, separation of biofilm and solitary bacteria by filtration, and disintegration of biofilms by ultrasonication. The filters used for this technique were evaluated for their nonspecific retention rates of solitary bacteria and for the efficiency of filtration for different concentrations of solitary bacteria in the presence of biofilms and other particles. The lethality and efficiency of disintegration of the sonication conditions used here were also evaluated. Isolation and quantification of bacteria by this technique is based on use of culture media. However, oligonucleotide probes, sera, or epifluorescent stains could also be used for direct characterization of the biofilm and solitary bacteria in the suspensions generated by this technique. Preliminary results from estimates of biofilm abundance in phyllosphere communities show that bacteria in biofilms constitute between about 10 and 40% of the total bacterial population on broad-leaf endive and parsley leaves.     

Heterogeneity in the ribosomal family of the yeast Yarrowia lipolytica: genomic organization and segregation studies

The cloned r-DNA units of Yarrowia lipolytica [Van Heerikhuizen et al., 39 (1985) 213-222] and their restriction fragments have been used to probe blots of genomic DNA of this yeast. Wild-type and laboratory strains were shown to contain two-to-five types of repeated units, each strain displaying a specific pattern. By comparing their restriction patterns, we could localize the differences between units within their spacer region. Tetrad analysis strongly suggested a clustered organization of each type of repeat as well as the occurrence of meiotic exchanges within the r-DNA family. Chromosome loss was induced by benomyl and allowed to map several r-DNA clusters on the same chromosome. All those results indicate that the Y. lipolytica r-DNA gene family is quite different from other yeasts.     

A comparison of extraction methods for the isolation of phospholipids from biological sources

Four classical methods, as well as a method presented in this paper, were compared as to their efficiency in extracting phospholipids from animal tissue. After the extractions, total lipids were separated quantitatively by DEAE-Sephadex chromatography into their acidic and nonacidic fractions. The two fractions were then further analyzed by gradient saturation high-performance thin-layer chromatography (HPTLC) combined with scanning photodensitometry after coloration with copper acetate. Of the five methods compared, the present and Christiansen's methods based upon single-phase solvent systems proved to be more efficient than biphasic extraction procedures. The undesirable discriminatory effect exhibited by biphasic solvent systems toward acidic phospholipids which were partly retained in the aqueous phase was confirmed by statistical evaluation of the HPTLC results. Total chromogenic response of acidic phospholipids extracted using biphasic solvent systems was shown to be lower by 10-35% in comparison to the single-phase method of Christiansen. The suitability of the present method for studies involving phospholipid synthesis was confirmed by monitoring the elimination of water-soluble compounds from the single-phase extracts using a classical phospholipid precursor, 2-[3H]glycerol-3-phosphate. The labeled compound was eliminated (99.3-100%) from the single-phase postcentrifugation supernatant, followed by DEAE-Sephadex chromatography.     

Randomised controlled trial of enalapril and beta blockers in non-diabetic chronic renal failure.

OBJECTIVE--To compare the ability of angiotensin converting enzyme inhibitors and beta blockers to slow the development of end stage renal failure in non-diabetic patients with chronic renal failure. DESIGN--Open randomised multicentre trial with three year follow up. SETTING--Outpatient departments of six French hospitals. PATIENTS--100 hypertensive patients with chronic renal failure (initial serum creatinine 200-400 mumol/l. 52 randomised to enalapril and 48 to beta blockers (conventional treatment). INTERVENTIONS--Enalapril or beta blocker was combined with frusemide and, if necessary, a calcium blocker or centrally acting drug in patients whose diastolic pressure remained above 90 mm Hg. RESULTS--17 patients receiving conventional treatment and 10 receiving enalapril developed end stage renal failure. The cumulative renal survival rate was significantly better in the enalapril group than in the conventional group (P < 0.05). The slope of the reciprocal serum creatinine concentration was steeper in the conventionally treated patients (-6.89 x 10(-5)l/mumol/month) than in the enalapril group (-4.17 x 10(-5)l/mumol/month; P < 0.05). No difference in blood pressure was found between groups. CONCLUSION--In hypertensive patients with chronic renal failure enalapril slows progression towards end stage renal failure compared with beta blockers. This effect was probably not mediated through controlling blood pressure.     

Tryptophanyl-tRNA synthetase is a major soluble protein species in bovine pancreas

Besides their central role in protein synthesis, aminoacyl-tRNA synthetases have been found or thought to be involved in other processes. We present here a study showing that tryptophanyl-tRNA synthetase has a surprising tissular distribution. Indeed, immunochemical determinations showed that in several bovine organs such as liver, kidney and heart, tryptophanyl-tRNA synthetase constitutes, as expected, about 0.02% of soluble proteins. In spleen, brain cortex, stomach, cerebellum or duodenum, this amount is about 10-times higher, and in pancreas it is 100-fold. There is no correlation between these amounts and the RNA content of the organs. Moreover, the concentration of another aminoacyl-tRNA synthetase (methionyl-tRNA synthetase) is higher in liver than in pancreas, while the amount of tRNATrp is not higher in pancreas than in liver as compared to other tRNAs. Among several interpretations, it is possible that tryptophanyl-tRNA synthetase is involved in a function other than tRNA aminoacylation. This unknown function would be specific to the differentiated organs, since fetal cerebellum and fetal pancreas contain the same amount of tryptophanyl-tRNA synthetase as adult liver.     

Corynebacterium glutamicum,a natural overproducer of succinic acid?

Corynebacterium glutamicum is well known as an important industrial amino acid producer. For a few years, its ability to produce organic acids, under micro‐aerobic or anaerobic conditions was demonstrated. This study is focused on the identification of the culture parameters influencing the organic acids production and, in particular, the succinate production, by this bacterium. Corynebacterium glutamicum 2262, used throughout this study, was a wild‐type strain, which was not genetically designed for the production of succinate. The oxygenation level and the residual glucose concentration appeared as two critical parameters for the organic acids production. The maximal succinate concentration (4.9 g L^?1) corresponded to the lower k_La value of 5 h^?1. Above 5 h^?1, a transient accumulation of the succinate was observed. Interestingly, the stop in the succinate production was concomitant with a lower threshold glucose concentration of 9 g L^?1. Taking into account this threshold, a fed‐batch culture was performed to optimize the succinate production with C. glutamicum 2262. The results showed that this wild‐type strain was able to produce 93.6 g L^?1 of succinate, which is one of the highest concentration reported in the literature.     

For a few years more: reductions in plant diversity 70 years after the last fire in Mediterranean forests

Plant Ecology - Changes in community diversity and dynamics after fires in Mediterranean ecosystems are rarely investigated more than a few years after the fire even though pronounced changes can...