Evaluation of concrete structures by combining non-destructive testing methods (SENSO project)

The management and maintenance of the built heritage is one of the main interests of the owners of concrete structures. The engineers wish to obtain quantitative information about concrete properties and their variability. Non-destructive testing (NDT) is very popular in this context as it quickly provides relevant information on the integrity and evolution of the material, but several kinds of indicators representative of the concrete condition need to be evaluated. A French Project, named SENSO, aims to develop methods for the non-destructive evaluation of concrete based on a multi-techniques approach. Several families of techniques are concerned (ultrasonic, electromagnetic, electrical, etc.). The main objective is to define the sensitivity of the techniques and the variability of the evaluation for each indicator concerned. To achieve this, a large experimental programme, involving a representative range of concretes and several indicators, has been carried out. A large database, linking the NDT observables and the indicators, allows the different observables to be distinguished in terms of quality (linked to the variability) and in terms of relevance for the characterisation of each indicator. The improvement of the indicator evaluation by means of technique combinatio

A high‐resolution view of the coordination environment in a paramagnetic metalloprotein from its magnetic properties

Metalloproteins constitute a significant fraction of the proteome of all organisms and their characterization is critical for both basic sciences and biomedical applications. A large portion of metalloproteins bind paramagnetic metal ions, and paramagnetic NMR spectroscopy has been widely used in their structural characterization. However, the signals of nuclei in the immediate vicinity of the metal center are often broadened beyond detection. In this work, we show that it is possible to determine the coordination environment of the paramagnetic metal in the protein at a resolution inaccessible to other techniques. Taking the structure of a diamagnetic analogue as a starting point, a geometry optimization is carried out by fitting the pseudocontact shifts obtained from first principles quantum chemical calculations to the experimental ones

Exploiting Mitochondrial Dysfunction for Effective Elimination of Imatinib-Resistant Leukemic Cells

Challenges today concern chronic myeloid leukemia (CML) patients resistant to imatinib. There is growing evidence that imatinib-resistant leukemic cells present abnormal glucose metabolism but the impact on mitochondria has been neglected. Our work aimed to better understand and exploit the metabolic alterations of imatinib-resistant leukemic cells. Imatinib-resistant cells presented high glycolysis as compared to sensitive cells. Consistently, expression of key glycolytic enzymes, at least partly mediated by HIF-1α, was modified in imatinib-resistant cells suggesting that imatinib-resistant cells uncouple glycolytic flux from pyruvate oxidation. Interestingly, mitochondria of imatinib-resistant cells exhibited accumulation of TCA cycle intermediates, increased NADH and low oxygen consumption. These mitochondrial alterations due to the partial failure of ETC were further confirmed in leukemic cells isolated from some imatinib-resistant CML patients. As a consequence, mitochondria generated more ROS than those of imatinib-sensitive cells. This, in turn, resulted in increased death of imatinib-resistant leukemic cells following in vitro or in vivo treatment with the pro-oxidants, PEITC and Trisenox, in a syngeneic mouse tumor model. Conversely, inhibition of glycolysis caused derepression of respiration leading to lower cellular ROS. In conclusion, these findings indicate that imatinib-resistant leukemic cells have an unexpected mitochondrial dysfunction that could be exploited for selective therapeutic intervention

NMR methods to monitor the enzymatic depolymerization of heparin

Heparin and the related glycosaminoglycan, heparan sulfate, are polydisperse linear polysaccharides that mediate numerous biological processes due to their interaction with proteins. Because of the structural complexity and heterogeneity of heparin and heparan sulfate, digestion to produce smaller oligosaccharides is commonly performed prior to separation and analysis. Current techniques used to monitor the extent of heparin depolymerization include UV absorption to follow product formation and size exclusion or strong anion exchange chromatography to monitor the size distribution of the components in the digest solution. In this study, we used 1H nuclear magnetic resonance (NMR) survey spectra and NMR diffusion experiments in conjunction with UV absorption measurements to monitor heparin depolymerization using the enzyme heparinase I. Diffusion NMR does not require the physical separation of the components in the reaction mixture and instead can be used to monitor the reaction solution directly in the NMR tube. Using diffusion NMR, the enzymatic reaction can be stopped at the desired time point, maximizing the abundance of larger oligosaccharides for protein-binding studies or completion of the reaction if the goal of the study is exhaustive digestion for characterization of the disaccharide composition. In this study, porcine intestinal mucosa heparin was depolymerized using the enzyme heparinase I. The unsaturated bond formed by enzymatic cleavage serves as a UV chromophore that can be used to monitor the progress of the depolymerization and for the detection and quantification of oligosaccharides in subsequent separations. The double bond also introduces a unique multiplet with peaks at 5.973, 5.981, 5.990, and 5.998 ppm in the 1H-NMR spectrum downfield of the anomeric region. This multiplet is produced by the proton of the C-4 double bond of the non-reducing end uronic acid at the cleavage site. Changes in this resonance were used to monitor the progression of the enzymatic digestion and compared to the profile obtained from UV absorbance measurements. In addition, in situ NMR diffusion measurements were explored for their ability to profile the different-sized components generated over the course of the digestion

Structure of a Wbl protein and implications for NO sensing by M. tuberculosis

Mycobacterium tuberculosis causes pulmonary tuberculosis (TB) and claims ~1.8 million human lives per annum. Host nitric oxide (NO) is important in controlling TB infection. M. tuberculosis WhiB1 is a NO-responsive Wbl protein (actinobacterial iron-sulfur proteins first identified in the 1970s). Until now, the structure of a Wbl protein has not been available. Here a NMR structural model of WhiB1 reveals that Wbl proteins are four-helix bundles with a core of three α-helices held together by a [4Fe-4S] cluster. The iron-sulfur cluster is required for formation of a complex with the major sigma factor (σA) and reaction with NO disassembles this complex. The WhiB1 structure suggests that loss of the iron-sulfur cluster (by nitrosylation) permits positively charged residues in the C-terminal helix to engage in DNA binding, triggering a major reprogramming of gene expression that includes components of the virulence-critical ESX-1 secretion system