Folding mechanisms steer the amyloid fibril formation propensity of highly homologous proteins

Significant advances in the understanding of the molecular determinants of fibrillogenesis can be expected from comparative studies of the aggregation propensities of proteins with highly homologous structures but different folding pathways. Here, we fully characterize, by means of stopped-flow, T-jump, CD and DSC experiments, the unfolding mechanisms of three highly homologous proteins, zinc binding Ros87 and Ml153-149 and zinc-lacking Ml452-151. The results indicate that the three proteins significantly differ in terms of stability and (un)folding mechanisms. Particularly, Ros87 and Ml153-149 appear to be much more stable to guanidine denaturation and are characterized by folding mechanisms including the presence of an intermediate. On the other hand, metal lacking Ml452-151 folds according to a classic two-state model. Successively, we have monitored the capabilities of Ros87, Ml452-151 and Ml153-149 to form amyloid fibrils under native conditions. Particularly, we show, by CD, fluorescence, DLS, TEM and SEM experiments, that after 168 hours, amyloid formation of Ros87 has started, while Ml153-149 has formed only amorphous aggregates and Ml452-151 is still monomeric in solution. This study shows how metal binding can influence protein folding pathways and thereby control conformational accessibility to aggregation-prone states, which in turn changes aggregation kinetics, shedding light on the role of metal ions in the development of protein deposition diseases

Ml proteins from Mesorhizobium loti and MucR from Brucella abortus: an AT-rich core DNA-target site and oligomerization ability

Mesorhizobium loti contains ten genes coding for proteins sharing high amino acid sequence identity with members of the Ros/MucR transcription factor family. Five of these Ros/MucR family members from Mesorhizobium loti (Ml proteins) have been recently structurally and functionally characterized demonstrating that Ml proteins are DNA-binding proteins. However, the DNA-binding studies were performed using the Ros DNA-binding site with the Ml proteins. Currently, there is no evidence as to when the Ml proteins are expressed during the Mesorhizobium loti life cycle as well as no information concerning their natural DNA-binding site. In this study, we examine the ml genes expression profile in Mesorhizobium loti and show that ml1, ml2, ml3 and ml5 are expressed during planktonic growth and in biofilms. DNA-binding experiments show that the Ml proteins studied bind a conserved AT-rich site in the promoter region of the exoY gene from Mesorhizobium loti and that the proteins make important contacts with the minor groove of DNA. Moreover, we demonstrate that the Ml proteins studied form higher-order oligomers through their N-terminal region and that the same AT-rich site is recognized by MucR from Brucella abortus using a similar mechanism involving contacts with the minor groove of DNA and oligomerization

Effect of biochar addition on soil microbial community in a wheat crop

Biochar is known to enhance soil fertility and C sequestration, but relatively little information is currently available about its effect on soil microbial community, a component of terrestrial ecosystems that plays a key role in nutrient cycling. This study tested the effects of soil amendment with two loads of wood-derived biochar (30 and 60tha-1) in a wheat crop in Tuscany (Italy). Soil samples were collected 3 and 14 months after treatments over two successive growing seasons, and analysed for pH, total organic C (Corg), extractable C (Cext), microbial biomass-C (Cmic), 25 specific microbial activities, mean substrate-induced respiration (mSIR) for 25 substrates, functional microbial diversity and bacterial genetic diversity. No significant effect of biochar treatment was observed on Corg, Cext, Cmic, microbial quotient (Cmic % Corg) or genetic diversity. An increase in mSIR, some specific microbial activities and soil pH, and a significant change in functional diversity were observed 3 months after treatment. In contrast, no effect of biochar was detected 14 months after treatment for the parameters considered, except for a small but significant increase in pH. Our data suggest that biochar addition stimulated soil microbial activity without causing any apparent disturbance, but this positive effect was very short-lived. © 2013

Zinc Fingers

Zinc finger (ZF) domains, that represent the majority of the DNA-binding motifs in eukaryotes, are involved in several processes ranging from RNA packaging to transcriptional activation, regulation of apoptosis, protein folding and assembly, and lipid binding. While their amino acid composition varies from one domain to the other, a shared feature is the coordination of a zinc ion, with a structural role, by a different combination of cysteines and histidines. The classical zinc finger domain (also called Cys2His2) that represents the most common class, uses two cysteines and two histidines to coordinate the metal ion, and forms a compact ββα architecture consisting in a β-sheet and an α-helix. GAG-knuckle resembles the classical ZF, treble clef and zinc ribbon are also well represented in the human genome. Zinc fingers are also present in prokaryotes. The first prokaryotic ZF domain found in the transcriptional regulator Ros protein was identified in Agrobacterium tumefaciens. It shows a Cys2His2 metal ion coordination sphere and folds in a domain significantly larger than its eukaryotic counterpart arranged in a βββαα topology. Interestingly, this domain does not strictly require the metal ion coordination to achieve the functional fold. Here, we report what is known on the main classes of eukaryotic and prokarotic ZFs, focusing our attention to the role of the metal ion, the folding mechanism, and the DNA binding. The hypothesis of a horizontal gene transfer from prokaryotes to eukaryotes is also discussed

3D human body pose estimation by SuperQuadrics

This paper presents a method for 3D Human Body pose estimation. 3D real data of the searched object is acquired by a multi-camera system and segmented by a special preprocessing algorithm based on clothing analysis. The human body model is built by nine SuperQuadrics (SQ) with a-priori known anthropometric scaling and shape parameters. The pose is estimated hierarchically by RANSAC-object search with a least square fitting 3D point cloud to SQ models: at first the body, and then the limbs. The solution is verified by evaluating the matching score, i.e. the number of inliers corresponding to a-piori chosen distance threshold, and comparing this score with admissible inlier threshold for the body and limbs. This method can be used for 3D object recognition, localization and pose estimation of Human Body