A dynamic view of enzyme catalysis

Recent experimental advances have shown that enzymes are flexible molecules, and point to a direct link between dynamics and catalysis. Movements span a wide time range, from nano- to milli-seconds. In this paper we introduce two aspects of enzyme flexibility that are treated with two appropriate techniques. First, transition path sampling is used to obtain an unbiased picture of the transition state ensemble in chorismate mutase, as well as its local flexibility and the energy flow during the chemical step. Second, we consider the binding and release of substrates in L-rhamnulose-1-phosphate aldolase. We have calculated the normal modes of the enzyme with the elastic network model. The lowest frequency modes generate active site deformations that change the coordination number of the catalytic zinc ion. The coordination lability of zinc allows the binding and release of substrates. Substitution of zinc by magnesium blocks the exchange of ligands. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Purinergic and nitrergic junction potential in the human colon

The aim of the present work is to investigate a putative junction transmission [nitric oxide (NO) and ATP] in the human colon and to characterize the electrophysiological and mechanical responses that might explain different functions from both neurotransmitters. Muscle bath and microelectrode techniques were performed on human colonic circular muscle strips. The NO donor sodium nitroprusside (10 microM), but not the P2Y receptor agonist adenosine 5'-O-2-thiodiphosphate (10 microM), was able to cause a sustained relaxation. NG-nitro-L-arginine (L-NNA) (1 mM), a NO synthase inhibitor, but not 2'-deoxy-N6-methyl adenosine 3',5'-diphosphate tetraammonium salt (MRS 2179) (10 microM), a P2Y antagonist, increased spontaneous motility. Electrical field stimulation (EFS) at 1 Hz caused fast inhibitory junction potentials (fIJPs) and a relaxation sensitive to MRS 2179 (10 microM). EFS at higher frequencies (5 Hz) showed biphasic IJP with fast hyperpolarization sensitive to MRS 2179 followed by sustained hyperpolarization sensitive to L-NNA; both drugs were needed to fully block the EFS relaxation at 2 and 5 Hz. Two consecutive single pulses induced MRS 2179-sensitive fIJPs that showed a rundown. The rundown mechanism was not dependent on the degree of hyperpolarization and was present after incubation with L-NNA (1 mM), hexamethonium (100 microM), MRS 2179 (1 microM), and NF023 (10 microM). We concluded that single pulses elicit ATP release from enteric motor neurons that cause a fIJP and a transient relaxation that is difficult to maintain over time; also, NO is released at higher frequencies causing a sustained hyperpolarization and relaxation. These differences might be responsible for complementary mechanisms of relaxation being phasic (ATP) and tonic (NO).     

Micropatterning of single endothelial cell shape reveals a tight coupling between nuclear volume in G1 and proliferation

Shape-dependent local differentials in cell proliferation are considered to be a major driving mechanism of structuring processes in vivo, such as embryogenesis, wound healing, and angiogenesis. However, the specific biophysical signaling by which changes in cell shape contribute to cell cycle regulation remains poorly understood. Here, we describe our study of the roles of nuclear volume and cytoskeletal mechanics in mediating shape control of proliferation in single endothelial cells. Micropatterned adhesive islands were used to independently control cell spreading and elongation. We show that, irrespective of elongation, nuclear volume and apparent chromatin decondensation of cells in G1 systematically increased with cell spreading and highly correlated with DNA synthesis (percent of cells in the S phase). In contrast, cell elongation dramatically affected the organization of the actin cytoskeleton, markedly reduced both cytoskeletal stiffness (measured dorsally with atomic force microscopy) and contractility (measured ventrally with traction microscopy), and increased mechanical anisotropy, without affecting either DNA synthesis or nuclear volume. Our results reveal that the nuclear volume in G1 is predictive of the proliferative status of single endothelial cells within a population, whereas cell stiffness and contractility are not. These findings show that the effects of cell mechanics in shape control of proliferation are far more complex than a linear or straightforward relationship. Our data are consistent with a mechanism by which spreading of cells in G1 partially enhances proliferation by inducing nuclear swelling and decreasing chromatin condensation, thereby rendering DNA more accessible to the replication machinery.     

Ultrastructure of Early Secondary Embryogenesis by Multicellular and Unicellular Pathways in Cork Oak (Quercus suber L.)

Early cellular events during secondary embryogenesis were studiedin a cork oak recurrent embryogenic system in which embryosarise either in a multicellular budding pathway from a compactmass of proliferation or from isolated single cells in friablecallus. The compact mass of proliferation originated from theepidermal cells at the hypocotyl whose growth and convolutionwas characterized by a decrease in the nucleus/cytoplasm ratioand a marked increase in storage products. The transition fromthe compact mass to meristematic primordia occurred at the peripheryand was accompanied by cell dedifferentiation and a drasticreduction of storage products. Meristematic primordia evolvedto globular embryos by the organization of a protodermis andtwo internal centres. Microscope analysis of friable callusshowed an hypothetical sequence from single cells to aggregatesof a few cells, meristematic cell clusters and globular embryos.Single cells showed typical features of embryogenic cells suchas rich cytoplasm and a large number of starch grains and lipidbodies. A progressive cell dedifferentiation and a drastic reductionof storage products was observed when aggregates of a few cellsand meristematic cell clusters were compared. Progressive bipolarizationin large meristematic cell clusters initiated globular embryoformation. The comparison of both embryogenic pathways at theultrastructural level showed that subcellular changes followa similar sequential pattern, especially with regard to thestorage products. The possible role of plastid extrusions andmultivesicular bodies in the changing pattern of starch metabolismduring embryogenesis is discussed. Copyright 2001 Annals ofBotany Company Quercus suber L, cork oak, somatic embryogenesis, multicellular budding, friable callus, ultrastructural studies     

The ΔF508 mutation in cystic fibrosis patients of Southern Italy

Summary Fifty one independent cystic fibrosis (CF) families originating from a restricted area of Southern Italy (Campania) have been analyzed for KM19 and XV2c haplotypes and the ΔF508 mutation: 54% of the total CF chromosomes show the ΔF508 mutation. No significative correlations were obtained when clinical score, radiological score,Pseudomonas colonization, or clinical symptoms at presentation were matched with the presence or absence of the ΔF508 mutation.     

From Life Cycle Assessment to Life Cycle Management

Life cycle assessment (LCA) is a widely accepted methodology to support decision‐making processes in which one compares alternatives, and that helps prevent shifting of environmental burdens along the value chain or among impact categories. According to regulation in the European Union (EU), the movement of waste needs to be reduced and, if unavoidable, the environmental gain from a specific waste treatment option requiring transport must be larger than the losses arising from transport. The EU explicitly recommends the use of LCA or life cycle thinking for the formulation of new waste management plans. In the last two revisions of the Industrial Waste Management Programme of Catalonia (PROGRIC), the use of a life cycle thinking approach to waste policy was mandated. In this article we explain the process developed to arrive at practical life cycle management (LCM) from what started as an LCA project. LCM principles we have labeled the “3/3” principle or the “good enough is best” principle were found to be essential to obtain simplified models that are easy to understand for legislators and industries, useful in waste management regulation, and, ultimately, feasible. In this article, we present the four models of options for the management of waste solvent to be addressed under Catalan industrial waste management regulation. All involved actors concluded that the models are sufficiently robust, are easy to apply, and accomplish the aim of limiting the transport of waste outside Catalonia, according to the principles of proximity and sufficiency.     

Alcaloïdes steroidiques du Malouetia Brachyloba et du M. Heudelotii

Malouetine and funtumafrine C have been isolated from the leaves of Malouetia brachyloba, indicating that this species is not very different from M. bequaertiana. Five conanine derivatives have been extracted from the leaves of M. heudelotii, a species which does not contain any alkaloid with a quaternary ammonium function.     

Alien plants of coastal dune habitats in southern Spain

Alien plants from psammophilous ecosystems have been studied in southern Spain. Sixteen sites, which are included in the Andalusian Network of Protected Spaces, have been chosen along 1100 km of Andalusian coasts in order to verify the degree of abundance and influence of these species on different communities. Out of the 26 localized species, the increase of some invasive plants, such as Arctotheca calendula, Agave americana, A. sisalana, Cortaderia selloana, Lantana camara and Opuntia dillenii, affects the conservation of some natural coastal ecosystems.