PGRP-LB: An Inside View into the Mechanism of the Amidase Reaction

Peptidoglycan recognition proteins (PGRPs) are ubiquitous among animals and play pivotal functions in insect immunity. Non-catalytic PGRPs are involved in the activation of immune pathways by binding to the peptidoglycan (PGN), whereas amidase PGRPs are capable of cleaving the PGN into non-immunogenic compounds. Drosophila PGRP-LB belongs to the amidase PGRPs and downregulates the immune deficiency (IMD) pathway by cleaving meso-2,6-diaminopimelic (meso-DAP or DAP)-type PGN. While the recognition process is well analyzed for the non-catalytic PGRPs, little is known about the enzymatic mechanism for the amidase PGRPs, despite their essential function in immune homeostasis. Here, we analyzed the specific activity of different isoforms of Drosophila PGRP-LB towards various PGN substrates to understand their specificity and role in Drosophila immunity. We show that these isoforms have similar activity towards the different compounds. To analyze the mechanism of the amidase activity, we performed site directed mutagenesis and solved the X-ray structures of wild-type Drosophila PGRP-LB and its mutants, with one of these structures presenting a protein complexed with the tracheal cytotoxin (TCT), a muropeptide derived from the PGN. Only the Y78F mutation abolished the PGN cleavage while other mutations reduced the activity solely. Together, our findings suggest the dynamic role of the residue Y78 in the amidase mechanism by nucleophilic attack through a water molecule to the carbonyl group of the amide function destabilized by Zn²⁺.     

Novel TTLL5 Variants Associated with Cone-Rod Dystrophy and Early-Onset Severe Retinal Dystrophy

Variants of the TTLL5 gene, which encodes tubulin tyrosine ligase-like family member five, are a rare cause of cone dystrophy (COD) or cone-rod dystrophy (CORD). To date, only a few TTLL5 patients have been clinically and genetically described. In this study, we report five patients harbouring biallelic variants of TTLL5. Four adult patients presented either COD or CORD with onset in the late teenage years. The youngest patient had a phenotype of early onset severe retinal dystrophy (EOSRD). Genetic analysis was performed by targeted next generation sequencing of gene panels and assessment of copy number variants (CNV). We identified eight variants, of which six were novel, including two large multiexon deletions in patients with COD or CORD, while the EOSRD patient harboured the novel homozygous p.(Trp640*) variant and three distinct USH2A variants, which might explain the observed rod involvement. Our study highlights the role of TTLL5 in COD/CORD and the importance of large deletions. These findings suggest that COD or CORD patients lacking variants in known genes may harbour CNVs to be discovered in TTLL5, previously undetected by classical sequencing methods. In addition, variable phenotypes in TTLL5-associated patients might be due to the presence of additional gene defects.     

Recognizing the importance of scale in the ecology and management of riverine fish

Processes affecting fish populations range in scale from local to global. Fish response is also scale‐dependent, with some activities varying locally while others depend on large‐scale connectivity within or between watersheds. These issues are still only partly recognized, with large‐scale research often affected by non‐independent sampling, weak inference, poor model testing or model over‐extrapolation. Available multi‐scale studies can reach different conclusions about factors affecting fishes from local studies, but results vary between examples. Potential explanations are (i) effects on fishes are context‐dependent; (ii) different species or life‐stages are limited in different ways; (iii) multi‐scale studies are too few for generalization. We advocate improved use of geostatistical tools to guide sampling or interpret the spatial extent of management problems, and we illustrate this using brown trout in Welsh streams. Our strongest recommendation is that fish ecologists recognize the importance of interactions across scales in quantifying effects on fishes so that management decisions can be better based on evidence rather than judgement or extrapolation. Copyright © 2006 John Wiley & Sons, Ltd.     

Ability of Selected Lactic Acid Bacteria to Ferment a Pearl Millet–Soybean Slurry to Produce Gruels for Complementary Foods for Young Children

Abstract: To assess the ability of lactic acid bacteria to improve some nutritional characteristics of the pearl millet–soybean slurry to prepare complementary foods for young children in African countries, inoculation was performed using strains previously selected for their ability to hydrolyse starch, phytate, or α-galactooligosaccharides (α-GOS). For the sake of comparison with the action of a natural microflora, fermentation was also performed by back slopping inoculation, that is, with a sample obtained from spontaneously fermented traditional pearl millet slurry obtained from a small scale processing unit in Burkina Faso (Ouagadougou). Starter cultures thrived on the slurry as shown by counts on MRS agar, TTGE fingerprints, and fermentation patterns. The fermentation of precooked slurries inoculated by back slopping or with mixed cultures containing the amylolytic strain Lb. plantarum A6 enabled partial starch hydrolysis. Corresponding gruels had a suitable consistency for young child feeding at high dry matter content, and a high energy density: 88.7 ± 4.2 and 75.8 ± 5.1 kcal/100 g of sweetened gruel, for the gruels inoculated by back slopping or with Lb. plantarum A6, respectively. Unexpectedly, no decrease in phytates was observed in any of the experiments, suggesting the presence of one or more inhibitory compounds in soybean. Furthermore, preprocessing conditions before fermentation affect the carbohydrate composition of slurry and have a more profound effect than fermentation on the reduction of the α-GOS content. Practical Application: This research aims to investigate the capacity of selected lactic acid bacteria to produce in a bioprocess high energy density and nutritious gruels for complementary feeding of young children from an African cereal (pearl millet) mixed with soybean.     

L2F anemometry measurements of the inter-row flow field within a transonic axial compressor provided with curved windows

INTRODUCTIONMultibladerowcalculationscanbeperformedusingsteady3DNScodesprovidedthatassumptionsaremadeforthecouplingofbladerows.Thesocalledmixing--planeapproachneedsthetangentialaveragesofflowquantitiesattheinterfaceplanebetweenadjacentbladerows.Butth...     

A first quantitative XPS study of the surface films formed, by exposure to water, on Mg and on the Mg-Al intermetallics: Al3Mg2 and Mg17Al12

An XPS investigation was carried out of the surface films, formed by exposure to ultrapure water, on mechanically ground Mg and the two Mg-Al intermetallic compounds: Al₃Mg₂ and Mg₁₇Al₁₂. The mechanically ground Mg surface had a film of MgO at the Mg metal surface covered by a Mg(OH)₂ layer, formed by the reaction of the MgO with water vapour in the air. Upon immersion in ultrapure water, this film converted to a duplex film with an inner MgO layer next to the Mg metal and an external porous hydroxide layer. For both intermetallics, the XPS data is consistent with (i) preferential dissolution of Mg and (ii) a 10 nm thick film on the surface after immersion in ultrapure water; the film composition on Al₃Mg₂ was AlMg_1.4O_0.2(OH)_5.4 whilst on Mg₁₇Al₁₂ the composition was AlMg_2.5(OH)₈.     

Examination of wooden shelves used in the ripening of a raw milk smear cheese by FTIR spectroscopy

The surface of wooden shelves used in the ripening of a raw milk smear cheese was examined by attenuated total reflection Fourier transform infrared (ATR-FTIR) spectroscopy. FTIR spectra of zones of wooden shelves in contact with cheeses or not (control) were acquired. This acquisition was performed either directly after cheeses removal or after brushing with water to remove cheese compounds deposited onto wood surfaces. Their analysis revealed that the degree of similarity between the ATR-FTIR spectra of unused and cleaned by brushing wooden shelves is higher than with the spectra of non-cleaned surfaces. The spectra of non-cleaned zones where cheeses were ripened namely differed in the 920–1180 cm^?1 and in the 1485–1780 cm^?1 regions. These regions can be assigned to proteins and polysaccharides and thus might correspond to deposits of cheese rind surface as well as to the presence of microbial biofilms on wooden shelves. Similarities in these spectral regions with those of cheese rind support this hypothesis. These observations indicate that ATR-FTIR spectroscopy is a rapid and valuable analytical tool to directly investigate the global chemical composition of very thin films such as microbial biofilms or cheese rind deposits present on wooden shelves which are not accessible to standard chemical methods.     

Determination of fruit origin by using 26S rDNA fingerprinting of yeast communities by PCR–DGGE: preliminary application to Physalis fruits from Egypt

The determination of geographical origin is a demand of the traceability system of import–export food products. One hypothesis for tracing the source of a product is by global analysis of the microbial communities of the food and statistical linkage of this analysis to the geographical origin of the food. For this purpose, a molecular technique employing 26S rDNA profiles generated by PCR–DGGE was used to detect the variation in yeast community structures of three species of Physalis fruit (Physalis ixocarpa Brat, Physalis pubescens L, Physalis pruinosa L) from four Egyptian regions (Qalyoubia, Minufiya, Beheira and Alexandria Governments). When the 26S rDNA profiles were analysed by multivariate analysis, distinct microbial communities were detected. The band profiles of Physalis yeasts from different Governments were specific for each location and could be used as a bar code to discriminate the origin of the fruits. This method is a new traceability tool which provides fruit products with a unique biological bar code and makes it possible to trace back the fruits to their original location. Copyright © 2009 John Wiley & Sons, Ltd.