New aspects on the utilization of inorganic sulphate during dentin formation

Summary Single, large doses of ³⁵S-labelled sulphate were administered intravenously to 5 female mice, 7 weeks of age. The animals were killed 10 and 30 min, 4 and 24 h, and 3 days after injection. Decalcified, epoxy-embedded specimens of the upper incisors and molars were subsequently studied by light and electron microscopic autoradiography. In dentin formation, the labelled sulphate passes via the odontoblasts into the predentin. From there, only a small portion enters the mineralised dentin, a large portion being unbound at the mineralising front.     

The scientific impact of the Structural Genomics Consortium: a protein family and ligand-centered approach to medically-relevant human proteins

As many of the structural genomics centers have ended their first phase of operation, it is a good point to evaluate the scientific impact of this endeavour. The Structural Genomics Consortium (SGC), operating from three centers across the Atlantic, investigates human proteins involved in disease processes and proteins from Plasmodium falciparum and related organisms. We present here some of the scientific output of the Oxford node of the SGC, where the target areas include protein kinases, phosphatases, oxidoreductases and other metabolic enzymes, as well as signal transduction proteins. The SGC has aimed to achieve extensive coverage of human gene families with a focus on protein–ligand interactions. The methods employed for effective protein expression, crystallization and structure determination by X-ray crystallography are summarized. In addition to the cumulative impact of accelerated delivery of protein structures, we demonstrate how family coverage, generic screening methodology, and the availability of abundant purified protein samples, allow a level of discovery that is difficult to achieve otherwise. The contribution of NMR to structure determination and protein characterization is discussed. To make this information available to a wide scientific audience, a new tool for disseminating annotated structural information was created that also represents an interactive platform allowing for a continuous update of the annotation by the scientific community.     

Circulating biomarkers of extracellular matrix remodeling and risk of atherosclerotic events

PURPOSE OF REVIEW: Disturbances of the synthesis and breakdown of the extracellular matrix of arterial walls have emerged as key features of the atherosclerotic process. Altered levels of circulating extracellular matrix markers have frequently been observed in relation to manifestations of atherosclerotic disease and its risk factors. RECENT FINDINGS: Research has been focused on the matrix-degrading metalloproteinases, their tissue inhibitors, and procollagen peptides. The most promising matrix metalloproteinase is matrix metalloproteinase-9, which has been observed to predict rapid coronary artery narrowing, ischemic heart disease incidence, abdominal aortic aneurysm expansion, worse outcome in stroke patients, and cardiovascular death. The use of tissue inhibitors of metalloproteinases for prognostication is uncertain thus far. The procollagen marker with most prognostic potential is the marker for type III collagen turnover rate, the N-terminal propeptide PIIINP, higher levels of which predict an adverse outcome after a myocardial infarction and in chronic heart failure, and portend abdominal aortic aneurysm expansion and risk of rupture. Also, the marker for type I collagen synthesis, the C-terminal propeptide PICP, predicts adverse outcomes following myocardial infarction and in chronic heart failure. Extracellular matrix remodeling is also a promising therapeutic target, being favorably affected by several conventional cardiovascular drugs and select dietary interventions. Synthetic matrix metalloproteinase inhibitors are also under development. SUMMARY: Circulating matrix markers have emerged as candidate biomarkers for predicting risk of subsequent atherosclerotic events. Future large longitudinal observational and intervention studies will determine the role of matrix biomarkers in diagnosis and prognostication, and as targets for intervention in cardiovascular diseases.     

Neuropeptide Y-family peptides and receptors in the elephant shark, Callorhinchus milii confirm gene duplications before the gnathostome radiation

We describe here the repertoire of neuropeptide Y (NPY) peptides and receptors in the elephant shark Callorhinchus milii, belonging to the chondrichthyans that diverged from the rest of the gnathostome (jawed vertebrate) lineage about 450 million years ago and the first chondrichthyan with a genome project. We have identified two peptide genes that are orthologous to NPY and PYY (peptide YY) in other vertebrates, and seven receptor genes orthologous to the Y1, Y2, Y4, Y5, Y6, Y7 and Y8 subtypes found in tetrapods and teleost fishes. The repertoire of peptides and receptors seems to reflect the ancestral configuration in the predecessor of all gnathostomes, whereas other lineages such as mammals and teleosts have lost one or more receptor genes or have acquired 1-2 additional peptide genes. Both the peptides and receptors showed broad and overlapping mRNA expression which may explain why some receptor gene losses could take place in some lineages, but leaves open the question why all the known ancestral receptors have been retained in the elephant shark.     

Differential expression analysis of Escherichia coli proteins using a novel software for relative quantitation of LC-MS/MS data

The study of changes in protein levels between samples derived from cells representing different biological conditions is a key to the understanding of cellular function. There are two main methods available that allow both for global scanning for significantly varying proteins and targeted profiling of proteins of interest. One method is based on 2-D gel electrophoresis and image analysis of labelled proteins. The other method is based on LC-MS/MS analysis of either unlabelled peptides or peptides derived from isotopically labelled proteins or peptides. In this study, the non-labelling approach was used involving a new software, DeCyder MS Differential Analysis Software (DeCyder MS) intended for automated detection and relative quantitation of unlabelled peptides in LC-MS/MS data.Total protein extracts of E. coli strains expressing varying levels of dihydrofolate reductase and integron integrase were digested with trypsin and analyzed using a nanoscale liquid chromatography system, Ettan MDLC, online connected to an LTQTM linear ion-trap mass spectrometer fitted with a nanospray interface. Acquired MS data were subjected to DeCyder MS analysis where 2-D representations of the peptide patterns from individual LC-MS/MS analyses were matched and compared.This approach to unlabelled quantitative analysis of the E. coli proteome resulted in relative protein abundances that were in good agreement with results obtained from traditional methods for measuring protein levels.     

In vitro self-assembly of the light harvesting pigment-protein LH2 revealed by ultrafast spectroscopy and electron microscopy

Controlled ensemble formation of protein-surfactant systems provides a fundamental concept for the realization of nanoscale devices with self-organizing capability. In this context, spectroscopic monitoring of pigment-containing proteins yields detailed structural information. Here we have studied the association behavior of the bacterial light-harvesting protein LH2 from Rhodobacter spheroides in an n,n-dimethyldodecylamine-n-oxide/water environment. Time-resolved studies of the excitation annihilation yielded information about aggregate sizes and packing of the protein complexes therein. The results are compared to transmission electron microscopy images of instantaneously frozen samples. Our data indicate the manifestation of different phases, which are discussed with respect to the thermodynamic equilibrium in ternary protein-surfactant-water systems. Accordingly, by varying the concentration the formation of different types of aggregates can be controlled. Conditions for the appearance of isolated LH2 complexes are defined.     

Inbreeding and sex-biased gene flow in the ant Formica exsecta

Abstract.— The objective of this study was to assess breeding and dispersal patterns of both males and females in a monogyne (a single queen per colony) population of ants. Monogyny is commonly associated with extensive nuptial flights, presumably leading to considerable gene flow over large areas. Opposite to these expectations we found evidence of both inbreeding and sex-biased gene flow in a monogyne population of Formica exsecta . We found a significant degree of population subdivision at a local scale (within islands) for queens (females heading established colonies) and workers, but not for colony fathers (the males mated to the colony queens). However, we found little evidence of population subdivision at a larger scale (among islands). More conclusive support for sex-biased gene flow comes from the analysis of isolation by distance on the largest island, and from assignment tests revealing differences in female and male philopatry. The genetic similarity between pairs of queens decreased significantly when geographical distance increased, demonstrating limited dispersal and isolation by distance in queens. By contrast, we found no such pattern for colony fathers. Furthermore, a significantly greater fraction of colony queens were assigned as having originated from the population of residence, as compared to colony fathers. Inbreeding coefficients were significantly positive for workers, but not for mother queens. The queen-male relatedness coefficient of 0.23 (regression relatedness) indicates that mating occurs between fairly close relatives. These results suggest that some monogyne species of ants have complex dispersal and mating systems that can result in genetic isolation by distance over small geographical scales. More generally, this study also highlights the importance of identifying the relevant scale in analyses of population structure and dispersal.