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171.
Initial steps in the synthesis of functional tRNAs require 5'- and 3'-processing of precursor tRNAs (pre-tRNAs), which in yeast mitochondria are achieved by two endonucleases, RNase P and RNase Z. In this study, using a combination of detergent-free Blue Native Gel Electrophoresis, proteomics and in vitro testing of pre-tRNA maturation, we reveal the physical association of these plus other mitochondrial activities in a large, stable complex of 136 proteins. It contains a total of seven proteins involved in RNA processing including RNase P and RNase Z, five out of six subunits of the mitochondrial RNA degradosome, components of the fatty acid synthesis pathway, translation, metabolism and protein folding. At the RNA level, there are the small and large rRNA subunits and RNase P RNA. Surprisingly, this complex is absent in an oar1Δ deletion mutant of the type II fatty acid synthesis pathway, supporting a recently published functional link between pre-tRNA processing and the FAS II pathway--apparently by integration into a large complex as we demonstrate here. Finally, the question of mt-RNase P localization within mitochondria was investigated, by GFP-tracing of a known protein subunit (Rpm2p). We find that about equal fractions of RNase P are soluble versus membrane-attached. 相似文献
172.
173.
Coutard B Danchin EG Oubelaid R Canard B Bignon C 《Protein expression and purification》2012,82(2):352-359
We previously reported the set up of an automated test for screening the refolding of recombinant proteins expressed as inclusion bodies in Escherichia coli[1]. The screen used 96 refolding buffers and was validated with 24 proteins, 70% of which remained soluble in at least one buffer. In the present paper, we have analyzed in more detail these experimental data to see if the refolding process can be driven by general rules. Notably, we found that proteins with an acidic isoelectric point (pI) refolded in buffers the average pH of which was alkaline and conversely. In addition, the number of refolding buffers wherein a protein remained soluble increased with the difference between its pI and the average pH of the buffers in which it refolded. A trend analysis of the other variables (ionic strength, detergents, etc.) was also performed. On the basis of this analysis, we devised and validated a new refolding screen made of a single buffer for acidic proteins and a single buffer for alkaline proteins. 相似文献
174.
175.
Isabelle Grechi Marie-Hélène Sauge Benoît Sauphanor Nadine Hilgert Rachid Senoussi & Françoise Lescourret 《Entomologia Experimentalis et Applicata》2008,128(3):369-379
Winter tree pruning is a cultural practice known to modify vegetative growth, which is likely to affect the development of pests. However, it has been poorly addressed as a cultural control method for diminishing the population levels of the green peach aphid, Myzus persicae (Sulzer) (Homoptera: Aphididae), in peach [ Prunus persica (L.) Batsch (Rosaceae)] orchards. In this study, we conducted a 2-year, on-station experiment to evaluate how winter pruning affects peach– M. persicae interactions, by examining tree vegetative growth, aphid population dynamics, and crop yield and fruit quality. We collected data under an insect-proof shelter on adult peach trees submitted to various levels of pruning and artificially infested with aphids. Our results showed that pruning enhanced shoot growth due to the proportion of growing shoots, which increased exponentially (10–60%), whereas the growth rate of growing shoots was not affected. The degree of infestation of peach trees increased with increasing pruning intensity. This effect was mainly due to the increase of the proportion of growing shoots, on which aphids developed better than on rosettes. In turn, the higher the aphid infestation, the higher the aphid-induced shoot-tip damage, leaf curling, and leaf fall that disturbed the growth of growing shoots. However, aphids did not considerably reduce fruit quality at harvest. They did not affect fresh fruit weight, and the refractometric index (indicator of sugar content) was reduced by only 3–4%. The relevance of winter pruning as a cultural method for pest control in orchards conducted under integrated fruit production guidelines is discussed. 相似文献
176.
Panchaud A Hansson J Affolter M Bel Rhlid R Piu S Moreillon P Kussmann M 《Molecular & cellular proteomics : MCP》2008,7(4):800-812
Identification and relative quantification of hundreds to thousands of proteins within complex biological samples have become realistic with the emergence of stable isotope labeling in combination with high throughput mass spectrometry. However, all current chemical approaches target a single amino acid functionality (most often lysine or cysteine) despite the fact that addressing two or more amino acid side chains would drastically increase quantifiable information as shown by in silico analysis in this study. Although the combination of existing approaches, e.g. ICAT with isotope-coded protein labeling, is analytically feasible, it implies high costs, and the combined application of two different chemistries (kits) may not be straightforward. Therefore, we describe here the development and validation of a new stable isotope-based quantitative proteomics approach, termed aniline benzoic acid labeling (ANIBAL), using a twin chemistry approach targeting two frequent amino acid functionalities, the carboxylic and amino groups. Two simple and inexpensive reagents, aniline and benzoic acid, in their (12)C and (13)C form with convenient mass peak spacing (6 Da) and without chromatographic discrimination or modification in fragmentation behavior, are used to modify carboxylic and amino groups at the protein level, resulting in an identical peptide bond-linked benzoyl modification for both reactions. The ANIBAL chemistry is simple and straightforward and is the first method that uses a (13)C-reagent for a general stable isotope labeling approach of carboxylic groups. In silico as well as in vitro analyses clearly revealed the increase in available quantifiable information using such a twin approach. ANIBAL was validated by means of model peptides and proteins with regard to the quality of the chemistry as well as the ionization behavior of the derivatized peptides. A milk fraction was used for dynamic range assessment of protein quantification, and a bacterial lysate was used for the evaluation of relative protein quantification in a complex sample in two different biological states. 相似文献
177.
The malaria parasite, Plasmodium falciparum, invades the red blood cells (RBCs) of its human host and initiates a series of morphological rearrangements within the host cell cytoplasm. The mature RBC has no endogenous trafficking machinery; therefore, the parasite generates novel structures to mediate protein transport. These include compartments called the Maurer's clefts (MC), which play an important role in the trafficking of parasite proteins to the surface of the host cell. Recent electron tomography studies have revealed MC as convoluted flotillas of flattened discs that are tethered to the RBC membrane, prompting speculation that the MC could, in one respect, represent an extracellular equivalent of the Golgi apparatus. Visualization of both resident and cargo proteins has helped decipher the signals and routes for trafficking of parasite proteins to the MC and beyond. 相似文献
178.
Cyril Ronco Antoine Millet Magali Plaisant Patricia Abbe Nedra Hamouda-Tekaya Stéphane Rocchi Rachid Benhida 《Bioorganic & medicinal chemistry letters》2017,27(10):2192-2196
We recently described a new family of bioactive molecules with interesting anti-cancer activities: the N-(4-(3-aminophenyl)thiazol-2-yl)acetamides. The lead compound of the series (1) displays significant anti-proliferative and cytotoxic activities against a panel of cancer cell lines, either sensitive or resistant to standard treatments. This molecule also shows a good pharmacological profile and high in vivo potency towards mice xenografts, without signs of toxicity on the animals. In the present article, we disclose the structure-activity relationships of this lead compound, which have provided clear information about the replacement of the acetamide function and the substitution pattern of the benzenesulfonamide ring. An improved high-yielding synthetic procedure towards these compounds has also been developed. Our drug design resulted in potency enhancement of 1, our new optimized lead compound being 19. These findings are of great interest to further improve this scaffold for the development of future clinical candidates. 相似文献
179.
Caio T. C. C. Rachid Fabiano C. Balieiro Eduardo S. Fonseca Raquel Silva Peixoto Guilherme M. Chaer James M. Tiedje Alexandre S. Rosado 《PloS one》2015,10(2)
Fungi are ubiquitous and important contributors to soil nutrient cycling, playing a vital role in C, N and P turnover, with many fungi having direct beneficial relationships with plants. However, the factors that modulate the soil fungal community are poorly understood. We studied the degree to which the composition of tree species affected the soil fungal community structure and diversity by pyrosequencing the 28S rRNA gene in soil DNA. We were also interested in whether intercropping (mixed plantation of two plant species) could be used to select fungal species. More than 50,000 high quality sequences were analyzed from three treatments: monoculture of Eucalyptus; monoculture of Acacia mangium; and a mixed plantation with both species sampled 2 and 3 years after planting. We found that the plant type had a major effect on the soil fungal community structure, with 75% of the sequences from the Eucalyptus soil belonging to Basidiomycota and 19% to Ascomycota, and the Acacia soil having a sequence distribution of 28% and 62%, respectively. The intercropping of Acacia mangium in a Eucalyptus plantation significantly increased the number of fungal genera and the diversity indices and introduced or increased the frequency of several genera that were not found in the monoculture cultivation samples. Our results suggest that management of soil fungi is possible by manipulating the composition of the plant community, and intercropped systems can be a means to achieve that. 相似文献
180.
Biofilm formation of Staphylococcus epidermidis on smooth polymer surfaces has been shown to be mediated by the ica operon. Upon activation of this operon, a polysaccharide intercellular adhesin (PIA) is synthesized that supports bacterial cell-to-cell contacts and triggers the production of thick, multilayered biofilms. Thus, the ica gene cluster represents a genetic determinant that significantly contributes to the virulence of specific Staphylococcus epidermidis strains. PIA synthesis has been reported recently to undergo a phase variation process. In this study, biofilm-forming Staphylococcus epidermidis strains and their PIA-negative phase variants were analysed genetically to investigate the molecular mechanisms of phase variation. We have characterized biofilm-negative variants by Southern hybridization with ica-specific probes, polymerase chain reaction and nucleotide sequencing. The data obtained in these analyses suggested that in approximately 30% of the variants the missing biofilm formation was due to the inactivation of either the icaA or the icaC gene by the insertion of the insertion sequence element IS256. Furthermore, it was shown that the transposition of IS256 into the ica operon is a reversible process. After repeated passages of the PIA-negative insertional mutants, the biofilm-forming phenotype could be restored. Nucleotide sequence analyses of the revertants confirmed the complete excision of IS256, including the initially duplicated 8 bp target sites. These results elucidate, for the first time, a molecular mechanism mediating phase variation in staphylcocci, and they demonstrate that a naturally occurring insertion sequence element is actively involved in the modulation of expression of a Staphylococcus virulence factor. 相似文献