In Silico screening for functional candidates amongst hypothetical proteins

Background  

The definition of a hypothetical protein is a protein that is predicted to be expressed from an open reading frame, but for which there is no experimental evidence of translation. Hypothetical proteins constitute a substantial fraction of proteomes of human as well as of other eukaryotes. With the general belief that the majority of hypothetical proteins are the product of pseudogenes, it is essential to have a tool with the ability of pinpointing the minority of hypothetical proteins with a high probability of being expressed.     

Preparation and characterization of the layered borophosphates M(H2O)2[B2P2O8(OH)2]·H2O (M = Fe, Co, Ni)

The isotypic layered transition metal borophosphates M^II(H₂O)₂[B₂P₂O₈(OH)₂]·H₂O (M^II = Fe, Co, Ni) were prepared under hydrothermal conditions. Their crystal structures were determined by single-crystal X-ray diffraction data and revealed an isotypic relationship to Mg(H₂O)₂[B₂P₂O₈(OH)₂]·H₂O, a structure containing wavy 6³ nets formed by tetrahedral phosphate and hydrogenborate groups interconnected in an alternating fashion by sharing common apices. The crystalline compounds were also characterized by chemical analyses, scanning electron microscopy, energy dispersive X-ray analyses, thermal analyses, IR-spectroscopy and magnetic susceptibility measurements.     

Proteomic analysis of iron acquisition,metabolic and regulatory responses of <Emphasis Type="Italic">Yersinia pestis</Emphasis> to iron starvation

Background  

The Gram-negative bacterium Yersinia pestis is the causative agent of the bubonic plague. Efficient iron acquisition systems are critical to the ability of Y. pestis to infect, spread and grow in mammalian hosts, because iron is sequestered and is considered part of the innate host immune defence against invading pathogens. We used a proteomic approach to determine expression changes of iron uptake systems and intracellular consequences of iron deficiency in the Y. pestis strain KIM6+ at two physiologically relevant temperatures (26°C and 37°C).     

X-ray crystal structure of leukocyte type core 2 beta1,6-N-acetylglucosaminyltransferase. Evidence for a convergence of metal ion-independent glycosyltransferase mechanism

Leukocyte type core 2 beta1,6-N-acetylglucosaminyltransferase (C2GnT-L) is a key enzyme in the biosynthesis of branched O-glycans. It is an inverting, metal ion-independent family 14 glycosyltransferase that catalyzes the formation of the core 2 O-glycan (Galbeta1-3[GlcNAcbeta1-6]GalNAc-O-Ser/Thr) from its donor and acceptor substrates, UDP-GlcNAc and the core 1 O-glycan (Galbeta1-3GalNAc-O-Ser/Thr), respectively. Reported here are the x-ray crystal structures of murine C2GnT-L in the absence and presence of the acceptor substrate Galbeta1-3GalNAc at 2.0 and 2.7A resolution, respectively. C2GnT-L was found to possess the GT-A fold; however, it lacks the characteristic metal ion binding DXD motif. The Galbeta1-3GalNAc complex defines the determinants of acceptor substrate binding and shows that Glu-320 corresponds to the structurally conserved catalytic base found in other inverting GT-A fold glycosyltransferases. Comparison of the C2GnT-L structure with that of other GT-A fold glycosyltransferases further suggests that Arg-378 and Lys-401 serve to electrostatically stabilize the nucleoside diphosphate leaving group, a role normally played by metal ion in GT-A structures. The use of basic amino acid side chains in this way is strikingly similar to that seen in a number of metal ion-independent GT-B fold glycosyltransferases and suggests a convergence of catalytic mechanism shared by both GT-A and GT-B fold glycosyltransferases.     

Antioxidative response mechanisms in halophytes: Their role in stress defence

Normal growth and development of plants is greatly dependent on the capacity to overcome environmental stresses. Environmental stress conditions like high salinity, drought, high incident light and low or high temperature cause major crop losses worldwide. A common denominator in all these adverse conditions is the production of reactive oxygen species (ROS) within different cellular compartments of the plant cell. Plants have developed robust mechanisms including enzymatic or nonenzymatic scavenging pathways to counter the deleterious effects of ROS production. There are a number of general reviews on oxidative stress in plants and few on the role of ROS scavengers during stress conditions. Here we review the regulation of antioxidant enzymes during salt stress in halophytes, especially mangroves. We conclude that (i) antioxidant enzymes protect halophytes from deleterious ROS production during salt stress, and (ii) genetic information from mangroves and other halophytes would be helpful in defining the roles of individual isoforms. This information would be critical in using the appropriate genes for oxidative stress defence for genetic engineering of enhanced stress tolerance in crop systems.     

The biology of habitat dominance; can microbes behave as weeds?

Competition between microbial species is a product of, yet can lead to a reduction in, the microbial diversity of specific habitats. Microbial habitats can resemble ecological battlefields where microbial cells struggle to dominate and/or annihilate each other and we explore the hypothesis that (like plant weeds) some microbes are genetically hard‐wired to behave in a vigorous and ecologically aggressive manner. These ‘microbial weeds’ are able to dominate the communities that develop in fertile but uncolonized – or at least partially vacant – habitats via traits enabling them to out‐grow competitors; robust tolerances to habitat‐relevant stress parameters and highly efficient energy‐generation systems; avoidance of or resistance to viral infection, predation and grazers; potent antimicrobial systems; and exceptional abilities to sequester and store resources. In addition, those associated with nutritionally complex habitats are extraordinarily versatile in their utilization of diverse substrates. Weed species typically deploy multiple types of antimicrobial including toxins; volatile organic compounds that act as either hydrophobic or highly chaotropic stressors; biosurfactants; organic acids; and moderately chaotropic solutes that are produced in bulk quantities (e.g. acetone, ethanol). Whereas ability to dominate communities is habitat‐specific we suggest that some microbial species are archetypal weeds including generalists such as: Pichia anomala, Acinetobacter spp. and Pseudomonas putida; specialists such as Dunaliella salina, Saccharomyces cerevisiae, Lactobacillus spp. and other lactic acid bacteria; freshwater autotrophs Gonyostomum semen and Microcystis aeruginosa; obligate anaerobes such as Clostridium acetobutylicum; facultative pathogens such as Rhodotorula mucilaginosa, Pantoea ananatis and Pseudomonas aeruginosa; and other extremotolerant and extremophilic microbes such as Aspergillus spp., Salinibacter ruber and Haloquadratum walsbyi. Some microbes, such as Escherichia coli, Mycobacterium smegmatis and Pseudoxylaria spp., exhibit characteristics of both weed and non‐weed species. We propose that the concept of nonweeds represents a ‘dustbin’ group that includes species such as Synodropsis spp., Polypaecilum pisce, Metschnikowia orientalis, Salmonella spp., and Caulobacter crescentus. We show that microbial weeds are conceptually distinct from plant weeds, microbial copiotrophs, r‐strategists, and other ecophysiological groups of microorganism. Microbial weed species are unlikely to emerge from stationary‐phase or other types of closed communities; it is open habitats that select for weed phenotypes. Specific characteristics that are common to diverse types of open habitat are identified, and implications of weed biology and open‐habitat ecology are discussed in the context of further studies needed in the fields of environmental and applied microbiology.     

The Rate of Decline in Small Fibre Function Assessed Using Axon Reflex-Mediated Neurogenic Vasodilatation and the Importance of Age Related Centile Values to Improve the Detection of Clinical Neuropathy

Background

The LDIflare technique (LDIflare) is a simple non-invasive test of small fibre function in dorsal foot skin involving skin heating and measuring the size of the resulting axon reflex-mediated vasodilator (flare) response using a laser Doppler imager (LDI). This study establishes age-related normative reference ranges for the test and determines the rate of decline in small fibre function per decade. Additionally, the potential value of using age related centiles rather than Receiver Operator Curves (ROC) was explored by comparison of the sensitivity and specificity of each analytic technique in identifying clinical neuropathy.

Methods

LDIflare areas were assessed in 94 healthy controls and 66 individuals with diabetes with (DN+, n = 31) and without clinical neuropathy (DN-, n = 35); neuropathy defined as a Neuropathy Disability Score ≥3. The age specific 5th centile values were used as the ‘cut-offs’ for the diagnosis of neuropathy from which sensitivity and specificity were calculated.

Results

There was a significant age dependant decrease in LDIflare size (r = −0.42, p<0.0001) with no significant gender differences. The LDIflare size reduced 0.56 cm² per decade which gives a percentage reduction of approximately 5.5% per decade. Using the normative 5th centiles as the cut-offs, the technique had a sensitivity of 77%, specificity of 90%, positive predictive value of 82% and negative predictive value of 87%.The ROC analysis gave a threshold of <3.66 cm² for the cut-off, resulting in a sensitivity of 75%, specificity of 85%, positive predictive value of 74% and negative predictive value of 86%.

Conclusions

There is an age dependent decrease in small fibre function in the foot of 5.5% per decade. Both analytic techniques demonstrate good sensitivity and specificity for detecting clinical neuropathy but the technique based on age centiles offers better diagnostic accuracy and is therefore proposed as the method of choice.