2-D structure of the Neurospora crassa plasma membrane ATPase as determined by electron cryomicroscopy.

Large, well-ordered 2-D crystals of the dodecylmaltoside complex of the Neurospora crassa plasma membrane H(+)-ATPase grow rapidly on the surface of a polyethylene glycol-containing mixture similar to that originally developed for growing 3-D crystals of this integral membrane transport protein. Negative stain electron microscopy of the crystals shows that many are single layers. Cryoelectron microscopy of unstained specimens indicates that the crystals have a p6 layer group with unit cell dimensions of a = b = 167 A. Image processing of selected electron micrographs has yielded a projection map at 10.3 A resolution. The repeating unit of the ATPase crystals comprises six 100 kDa ATPase monomers arranged in a symmetrical ring. The individual monomers in projection are shaped like a boot. These results provide the first indications of the molecular structure of the H(+)-ATPase molecule. They also establish the feasibility of precipitant-induced surface growth as a rapid, simple alternative to conventional methods for obtaining 2-D crystals of the integral membrane proteins useful for structure analysis.     

Characterization of the TRBP domain required for Dicer interaction and function in RNA interference

Background  

Dicer, Ago2 and TRBP are the minimum components of the human RNA-induced silencing complex (RISC). While Dicer and Ago2 are RNases, TRBP is the double-stranded RNA binding protein (dsRBP) that loads small interfering RNA into the RISC. TRBP binds directly to Dicer through its C-terminal domain.     

Genetic variation in the effect of a facultative symbiont on host-plant use by pea aphids

Ecological specialisation on different host plants occurs frequently among phytophagous insects and is normally assumed to have a genetic basis. However, insects often carry microbial symbionts, which may play a role in the evolution of specialisation. The bacterium Regiella insecticola is a facultative symbiont of pea aphids (Acyrthosiphon pisum) where it is found most frequently in aphid clones feeding on Trifolium giving rise to the hypothesis that it may improve aphid performance on this plant. A study in which R. insecticola was eliminated from a single naturally infected aphid clone supported the hypothesis, but a second involving two aphid clones did not find the same effect. We created a series of new pea aphid–R. insecticola associations by injecting different strains of bacteria into five aphid clones uninfected by symbionts. For all aphid clones, the bacteria decreased the rate at which aphids accepted Vicia faba as a food plant and reduced performance on this plant. Their effect on aphids given Trifolium pratense was more complex: R. insecticola negatively affected acceptance by all aphid clones, had no effect on the performance of four aphid clones, but increased performance of a fifth, thus demonstrating genetic variation in the effect of R. insecticola on pea aphid host use. We discuss how these results may explain the distribution and frequency of this symbiont across different aphid populations. Julia Ferrari and Claire L. Scarborough contributed equally to the work.     

Expression and secretion of biologically active human atrial natriuretic peptide in Saccharomyces cerevisiae

A hybrid gene was constructed containing a fusion between the DNA sequences encoding the secretory precursor of the yeast mating pheromone alpha-factor and a synthetic sequence encoding a biologically active 24-amino acid carboxyl-terminal portion of the human atrial natriuretic peptide (hANP) precursor. Transformation of Saccharomyces cerevisiae with the hybrid gene resulted in the yeast cells secreting biologically active hANP into the extracellular medium. The secreted hANP was purified and found to be accurately processed at the junction in the chimeric alpha-factor/hANP protein, producing the desired mature hANP amino terminus. The secreted product was also folded correctly with respect to the single disulfide bond. However, the carboxyl terminus of the secreted hANP material was heterogeneous such that the major form lacked the last two amino acids of the peptide while the minor form was the full length material. The observed processing at the carboxyl terminus of the secreted hANP may reflect a normal processing event involved in alpha-factor peptide maturation.     

Patterns and Emerging Trends in Global Ocean Health

International and regional policies aimed at managing ocean ecosystem health need quantitative and comprehensive indices to synthesize information from a variety of sources, consistently measure progress, and communicate with key constituencies and the public. Here we present the second annual global assessment of the Ocean Health Index, reporting current scores and annual changes since 2012, recalculated using updated methods and data based on the best available science, for 221 coastal countries and territories. The Index measures performance of ten societal goals for healthy oceans on a quantitative scale of increasing health from 0 to 100, and combines these scores into a single Index score, for each country and globally. The global Index score improved one point (from 67 to 68), while many country-level Index and goal scores had larger changes. Per-country Index scores ranged from 41–95 and, on average, improved by 0.06 points (range -8 to +12). Globally, average scores increased for individual goals by as much as 6.5 points (coastal economies) and decreased by as much as 1.2 points (natural products). Annual updates of the Index, even when not all input data have been updated, provide valuable information to scientists, policy makers, and resource managers because patterns and trends can emerge from the data that have been updated. Changes of even a few points indicate potential successes (when scores increase) that merit recognition, or concerns (when scores decrease) that may require mitigative action, with changes of more than 10–20 points representing large shifts that deserve greater attention. Goal scores showed remarkably little covariance across regions, indicating low redundancy in the Index, such that each goal delivers information about a different facet of ocean health. Together these scores provide a snapshot of global ocean health and suggest where countries have made progress and where a need for further improvement exists.     

Design, synthesis and structure-activity relationship of a series of arginine aldehyde factor Xa inhibitors. Part 1: structures based on the (D)-Arg-Gly-Arg tripeptide sequence

A series of arginine aldehyde inhibitors was designed as transition state (TS) analogues based on the known factor Xa specific substrate Cbz-D-Arg-Gly-Arg-pNA. BnSO2-(D)Arg-Gly-Arg-H (20) was found to be the most potent and selective inhibitor of factor Xa and prothrombinase activity in this series.