1H and 15N nuclear magnetic resonance assignment and secondary structure of the cytotoxic ribonuclease alpha-Sarcin.

The ribosome-inactivating protein alpha-Sarcin (alpha S) is a 150-residue fungal ribonuclease that, after entering sensitive cells, selectively cleaves a single phosphodiester bond in an universally conserved sequence of the major rRNA to inactivate the ribosome and thus exert its cytotoxic action. As a first step toward establishing the structure-dynamics-function relationships in this system, we have carried out the assignment of the 1H and 15N NMR spectrum of alpha S on the basis of homonuclear (1H-1H) and heteronuclear (1H-15N) two-dimensional correlation spectra of a uniformly 15N-labeled sample, and two selectively 15N-labeled (Tyr and Phe) samples, as well as a single three-dimensional experiment. The secondary structure of alpha S, as derived from the characteristic patterns of dipolar connectivities between backbone protons, conformational chemical shifts, and the protection of backbone amide protons against exchange, consists of a long N-terminal beta-hairpin, a short alpha-helical segment, and a C-terminal beta-sheet of five short strands arranged in a + 1, + 1, + 1, + 1 topology, connected by long loops in which the 13 Pro residues are located.     

Role of fibronectin in the adhesion of Friend erythroleukemia cells

Friend Erythroleukemia Cells (FLC) (745 A and FW clones), normally growing in suspension, tend to adhere to fibroblast monolayers, but not to epithelial cells. Co-cultivation of FLC with Human Embryo Fibroblasts (HEF) resulted in the selection of adhesive Friend Cells. After 16 subcultures, we were able to isolate clones of adhesive FLC that grow in monolayer on plastic tissue culture plates. Both the binding of FLC to fibroblasts and of the adhesive clones to the plastic surfaces is completely suppressed by fibronectin antiserum, thus suggesting that fibronectin is responsible for FLC adhesion. Adhesive FLC clones maintain the ability to differentiate upon induction by DMSO.     

Cell membrane is a more sensitive target than cytoplasm to dense ionization produced by auger electrons

To improve radioimmunotherapy with Auger electron emitters, we assessed whether the biological efficiency of (125)I varied according to its localization. A-431 and SK-OV-3 carcinoma cells were incubated with increasing activities (0-4 MBq/ml) of (125)I-labeled vectors targeting the cell membrane, the cytoplasm or the nucleus. We then measured cell survival by clonogenic assay and the mean radiation dose to the nucleus by assessing the cellular medical internal radiation dose (MIRD). The relationship between survival and the radiation dose delivered was investigated with a linear mixed regression model. For each cell line, we obtained dose-response curves for the three targets and the reference values (i.e., the dose leading to 75, 50 or 37% survival). When cell survival was expressed as a function of the total cumulative decays, nuclear (125)I disintegrations were more harmful than disintegrations in the cytoplasm or at the cell membrane. However, when survival was expressed as a function of the mean radiation dose to the nucleus, toxicity was significantly higher when (125)I was targeted to the cell membrane than to the cytoplasm. These findings indicate that the membrane is a more sensitive target than the cytoplasm for the dense ionization produced by Auger electrons. Moreover, cell membrane targeting is as cytotoxic as nuclear targeting in SK-OV-3 cells. We suggest that targeting the membrane rather than the cytoplasm may contribute to the development of more efficient radioimmunotherapies based on Auger electron radiation, also because most of the available vectors are directed against cell surface antigens.     

Exaggerated trans-membrane charge of ammonium transporters in nutrient-poor marine environments

Transporter proteins are a vital interface between cells and their environment. In nutrient-limited environments, microbes with transporters that are effective at bringing substrates into their cells will gain a competitive advantage over variants with reduced transport function. Microbial ammonium transporters (Amt) bring ammonium into the cytoplasm from the surrounding periplasm space, but diagnosing Amt adaptations to low nutrient environments solely from sequence data has been elusive. Here, we report altered Amt sequence amino acid distribution from deep marine samples compared to variants sampled from shallow water in two important microbial lineages of the marine water column community—Marine Group I Archaea (Thermoproteota) and the uncultivated gammaproteobacterial lineage SAR86. This pattern indicates an evolutionary pressure towards an increasing dipole in Amt for these clades in deep ocean environments and is predicted to generate stronger electric fields facilitating ammonium acquisition. This pattern of increasing dipole charge with depth was not observed in lineages capable of accessing alternative nitrogen sources, including the abundant alphaproteobacterial clade SAR11. We speculate that competition for ammonium in the deep ocean drives transporter sequence evolution. The low concentration of ammonium in the deep ocean is therefore likely due to rapid uptake by Amts concurrent with decreasing nutrient flux.     

Morphological distance and inter-nest distance account for intra-specific prey overlap in digger wasps (Hymenoptera: Crabronidae)

Although inter-individual diet variation is common in predatory wasp populations, the factors accounting for such variation are still largely unknown. Here, we asked if paired diet dissimilarity in three species of digger wasps correlates with morphological distance and inter-nest distance, two factors previously linked to diet partitioning in vertebrates. Results sharply differed among species and generations. All sampled populations showed significant inter-individual diet variation for prey taxa, but only in half of the cases for prey size. In one generation of two species [Bembix zonata Klug and Stizus continuus (Klug)], similar-sized wasps had similar prey taxonomic spectra (and for S. continuus also similar prey size spectra), a phenomenon which probably reduces intra-specific competition. In addition, B. zonata females nesting closer to each other had more similar prey taxonomic spectra, suggesting that distant females probably hunt on different patches that harbour different prey species. For the females of a further species (Bembix merceti Parker), pairwise size difference and inter-nest distance did not affect prey dissimilarity. Both morphological distance and inter-nest distance are potentially important in shaping the overlap of individual resource use in wasps, though probably only in certain conditions such as a highly clumped distribution of nests and size-related constraints on prey selection.     

Analysis of the Electronic Circular Dichroism Spectrum of (−)–[9](2,5)Pyridinophane

The results of a combined experimental and theoretical study of the electronic circular dichroism spectrum of (?)–[9] (2,5)Pyridinophane are presented. The features observed in the measured spectrum in the 180–350 nm wavelength region are reproduced by a series of calculations carried out within a density functional time‐dependent approach, coupled with a vibronic analysis allowing rationalizing unambiguously the chiral response of the molecule. Chirality 24:994–1004, 2012. © 2012 Wiley Periodicals, Inc.