The mechanisms of humic substances self-assembly with biological molecules: The case study of the prion protein

Humic substances (HS) are the largest constituent of soil organic matter and are considered as a key component of the terrestrial ecosystem. HS may facilitate the transport of organic and inorganic molecules, as well as the sorption interactions with environmentally relevant proteins such as prions. Prions enter the environment through shedding from live hosts, facilitating a sustained incidence of animal prion diseases such as Chronic Wasting Disease and scrapie in cervid and ovine populations, respectively. Changes in prion structure upon environmental exposure may be significant as they can affect prion infectivity and disease pathology. Despite its relevance, the mechanisms of prion interaction with HS are still not completely understood. The goal of this work is to advance a structural-level picture of the encapsulation of recombinant, non-infectious, prion protein (PrP) into different natural HS. We observed that PrP precipitation upon addition of HS is mainly driven by a mechanism of “salting-out” whereby PrP molecules are rapidly removed from the solution and aggregate in insoluble adducts with humic molecules. Importantly, this process does not alter the protein folding since insoluble PrP retains its α-helical content when in complex with HS. The observed ability of HS to promote PrP insolubilization without altering its secondary structure may have potential relevance in the context of “prion ecology”. These results suggest that soil organic matter interacts with prions possibly without altering the protein structures. This may facilitate prions preservation from biotic and abiotic degradation leading to their accumulation in the environment

A Complete Pathway Model for Lipid A Biosynthesis in Escherichia coli.

Lipid A is a highly conserved component of lipopolysaccharide (LPS), itself a major component of the outer membrane of Gram-negative bacteria. Lipid A is essential to cells and elicits a strong immune response from humans and other animals. We developed a quantitative model of the nine enzyme-catalyzed steps of Escherichia coli lipid A biosynthesis, drawing parameters from the experimental literature. This model accounts for biosynthesis regulation, which occurs through regulated degradation of the LpxC and WaaA (also called KdtA) enzymes. The LpxC degradation signal appears to arise from the lipid A disaccharide concentration, which we deduced from prior results, model results, and new LpxK overexpression results. The model agrees reasonably well with many experimental findings, including the lipid A production rate, the behaviors of mutants with defective LpxA enzymes, correlations between LpxC half-lives and cell generation times, and the effects of LpxK overexpression on LpxC concentrations. Its predictions also differ from some experimental results, which suggest modifications to the current understanding of the lipid A pathway, such as the possibility that LpxD can replace LpxA and that there may be metabolic channeling between LpxH and LpxB. The model shows that WaaA regulation may serve to regulate the lipid A production rate when the 3-deoxy-D-manno-oct-2-ulosonic acid (KDO) concentration is low and/or to control the number of KDO residues that get attached to lipid A. Computation of flux control coefficients showed that LpxC is the rate-limiting enzyme if pathway regulation is ignored, but that LpxK is the rate-limiting enzyme if pathway regulation is present, as it is in real cells. Control also shifts to other enzymes if the pathway substrate concentrations are not in excess. Based on these results, we suggest that LpxK may be a much better drug target than LpxC, which has been pursued most often

Rac and Rho GTPases in cancer cell motility control

Rho GTPases represent a family of small GTP-binding proteins involved in cell cytoskeleton organization, migration, transcription, and proliferation. A common theme of these processes is a dynamic reorganization of actin cytoskeleton which has now emerged as a major switch control mainly carried out by Rho and Rac GTPase subfamilies, playing an acknowledged role in adaptation of cell motility to the microenvironment. Cells exhibit three distinct modes of migration when invading the 3 D environment. Collective motility leads to movement of cohorts of cells which maintain the adherens junctions and move by photolytic degradation of matrix barriers. Single cell mesenchymal-type movement is characterized by an elongated cellular shape and again requires extracellular proteolysis and integrin engagement. In addition it depends on Rac1-mediated cell polarization and lamellipodia formation. Conversely, in amoeboid movement cells have a rounded morphology, the movement is independent from proteases but requires high Rho GTPase to drive elevated levels of actomyosin contractility. These two modes of cell movement are interconvertible and several moving cells, including tumor cells, show an high degree of plasticity in motility styles shifting ad hoc between mesenchymal or amoeboid movements. This review will focus on the role of Rac and Rho small GTPases in cell motility and in the complex relationship driving the reciprocal control between Rac and Rho granting for the opportunistic motile behaviour of aggressive cancer cells. In addition we analyse the role of these GTPases in cancer progression and metastatic dissemination

Consensus guidelines for the use and interpretation of angiogenesis assays

The formation of new blood vessels, or angiogenesis, is a complex process that plays important roles in growth and development, tissue and organ regeneration, as well as numerous pathological conditions. Angiogenesis undergoes multiple discrete steps that can be individually evaluated and quantified by a large number of bioassays. These independent assessments hold advantages but also have limitations. This article describes in vivo, ex vivo, and in vitro bioassays that are available for the evaluation of angiogenesis and highlights critical aspects that are relevant for their execution and proper interpretation. As such, this collaborative work is the first edition of consensus guidelines on angiogenesis bioassays to serve for current and future reference

Quantitative comparison of catalytic mechanisms and overall reactions in convergently evolved enzymes : implications for classification of enzyme function

The authors thank the National Institutes of Health (NIH R01 GM60595 to PCB) and the Scottish Universities Life Sciences Alliance (SULSA to JBOM) for funding.Functionally analogous enzymes are those that catalyze similar reactions on similar substrates but do not share common ancestry, providing a window on the different structural strategies nature has used to evolve required catalysts. Identification and use of this information to improve reaction classification and computational annotation of enzymes newly discovered in the genome projects would benefit from systematic determination of reaction similarities. Here, we quantified similarity in bond changes for overall reactions and catalytic mechanisms for 95 pairs of functionally analogous enzymes (non-homologous enzymes with identical first three numbers of their EC codes) from the MACiE database. Similarity of overall reactions was computed by comparing the sets of bond changes in the transformations from substrates to products. For similarity of mechanisms, sets of bond changes occurring in each mechanistic step were compared; these similarities were then used to guide global and local alignments of mechanistic steps. Using this metric, only 44% of pairs of functionally analogous enzymes in the dataset had significantly similar overall reactions. For these enzymes, convergence to the same mechanism occurred in 33% of cases, with most pairs having at least one identical mechanistic step. Using our metric, overall reaction similarity serves as an upper bound for mechanistic similarity in functional analogs. For example, the four carbon-oxygen lyases acting on phosphates (EC 4.2.3) show neither significant overall reaction similarity nor significant mechanistic similarity. By contrast, the three carboxylic-ester hydrolases (EC 3.1.1) catalyze overall reactions with identical bond changes and have converged to almost identical mechanisms. The large proportion of enzyme pairs that do not show significant overall reaction similarity (56%) suggests that at least for the functionally analogous enzymes studied here, more stringent criteria could be used to refine definitions of EC sub-subclasses for improved discrimination in their classification of enzyme reactions. The results also indicate that mechanistic convergence of reaction steps is widespread, suggesting that quantitative measurement of mechanistic similarity can inform approaches for functional annotation.Publisher PDFPeer reviewe

National greenhouse-gas accounting for effective climate policy on international trade

National greenhouse-gas accounting should reflect how countries’ policies and behaviours affect global emissions. Actions that contribute to reduced global emissions should be credited, and actions that increase them should be penalized. This is essential if accounting is to serve as accurate guidance for climate policy. Yet this principle is not satisfied by the two most common accounting methods. Production-based accounting used under the Kyoto Protocol does not account for carbon leakage — the phenomenon of countries reducing their domestic emissions by shifting carbon-intensive production abroad1. Consumption-based accounting2,3 (also called carbon footprinting) does not credit countries for cleaning up their export industries, and it also punishes some types of trade that could contribute to more carbon efficient production worldwide. We propose an improvement to consumption-based carbon accounting that takes technology differences in export sectors into account and thereby tends to more correctly reflect how national policy changes affect total global emissions. We also present empirical results showing how this new measure redraws the global emissions map