Community-wide assessment of protein-interface modeling suggests improvements to design methodology

The CAPRI (Critical Assessment of Predicted Interactions) and CASP (Critical Assessment of protein Structure Prediction) experiments have demonstrated the power of community-wide tests of methodology in assessing the current state of the art and spurring progress in the very challenging areas of protein docking and structure prediction. We sought to bring the power of community-wide experiments to bear on a very challenging protein design problem that provides a complementary but equally fundamental test of current understanding of protein-binding thermodynamics. We have generated a number of designed protein-protein interfaces with very favorable computed binding energies but which do not appear to be formed in experiments, suggesting that there may be important physical chemistry missing in the energy calculations. A total of 28 research groups took up the challenge of determining what is missing: we provided structures of 87 designed complexes and 120 naturally occurring complexes and asked participants to identify energetic contributions and/or structural features that distinguish between the two sets. The community found that electrostatics and solvation terms partially distinguish the designs from the natural complexes, largely due to the nonpolar character of the designed interactions. Beyond this polarity difference, the community found that the designed binding surfaces were, on average, structurally less embedded in the designed monomers, suggesting that backbone conformational rigidity at the designed surface is important for realization of the designed function. These results can be used to improve computational design strategies, but there is still much to be learned; for example, one designed complex, which does form in experiments, was classified by all metrics as a nonbinder.     

Growth dynamics of non-toxic Pseudo-nitzschia delicatissima and toxic P. seriata (Bacillariophyceae) under simulated spring and summer photoperiods

Marine planktonic diatoms of the genus Pseudo-nitzschia Peragallo have been responsible for amnesic shellfish poisoning (ASP) events worldwide through the production of the neurotoxin domoic acid (DA). The appearance and toxicity of Pseudo-nitzschia species is variable throughout the year and potentially linked to changes in environmental parameters; many ASP events occur in relatively high latitudes where day length is particularly variable with season. In UK waters, shellfish monitoring has prevented any impact on human health but has led to long-term closures of fisheries, with severe economic consequences. Laboratory experiments on two Pseudo-nitzschia species typically found in Scottish West Coast waters during spring (short photoperiod (SP)) and summer (long photoperiod (LP)) conditions were conducted to determine the influence of photoperiod on their growth and toxicity. Results indicated that non-toxic P. delicatissima (Cleve) Heiden achieved a greater cell density under SP (9-h light:15-h dark (L:D) cycle). For toxin-producing P. seriata (Cleve) H. Peragallo, a LP (18-h L:6-h D cycle) resulted in an enhanced growth rate, cell yield and total toxin production, but it decreased the toxin production per cell. A better understanding of the response of Pseudo-nitzschia species to photoperiod and other foreseeable environmental variables may help predict the appearance of toxic strains.     

Interrupter resistance elucidated by alveolar pressure measurement in open-chest normal dogs

The interrupter method for measuring respiratory system resistance involves rapidly interrupting flow at the mouth while measuring the pressure just distal to the point of interruption. The pressure signal observed invariably exhibits two distinct phases. The first phase is a very rapid jump, designated delta Pinit, which occurs immediately on interruption of flow. The second phase is designated delta Pdif and is a further pressure change in the same direction as delta Pinit but evolving over several seconds. The physiological interpretations of delta Pinit and delta Pdif have been somewhat unclear. Delta Pinit has been taken to equal the pressure drop across the pulmonary airways, possibly with a contribution from the tissues of the respiratory system. Delta Pdif can arise, in principle, from two sources: gas redistribution throughout the lung after interruption of flow and stress recovery within the tissues. To resolve these issues we performed interruption experiments on anesthetized paralyzed, tracheotomized, open-chest normal dogs during passive expiration while measuring alveolar pressures at three sites with alveolar capsules. We found that, in the absence of the chest wall, delta Pinit reflects only the resistance of the airways and that delta Pdif can be ascribed almost entirely to the stress recovery properties of lung tissues.     

Examining the global distribution of dominant archaeal populations in soil

Archaea, primarily Crenarchaeota, are common in soil; however, the structure of soil archaeal communities and the factors regulating their diversity and abundance remain poorly understood. Here, we used barcoded pyrosequencing to comprehensively survey archaeal and bacterial communities in 146 soils, representing a multitude of soil and ecosystem types from across the globe. Relative archaeal abundance, the percentage of all 16S rRNA gene sequences recovered that were archaeal, averaged 2% across all soils and ranged from 0% to >10% in individual soils. Soil C:N ratio was the only factor consistently correlated with archaeal relative abundances, being higher in soils with lower C:N ratios. Soil archaea communities were dominated by just two phylotypes from a constrained clade within the Crenarchaeota, which together accounted for >70% of all archaeal sequences obtained in the survey. As one of these phylotypes was closely related to a previously identified putative ammonia oxidizer, we sampled from two long-term nitrogen (N) addition experiments to determine if this taxon responds to experimental manipulations of N availability. Contrary to expectations, the abundance of this dominant taxon, as well as archaea overall, tended to decline with increasing N. This trend was coupled with a concurrent increase in known N-oxidizing bacteria, suggesting competitive interactions between these groups.     

The projected effects of climatic and vegetation changes on the distribution and diversity of Southeast Asian bats

Southeast‐Asia (SEA) constitutes a global biodiversity hotspot, but is exposed to extensive deforestation and faces numerous threats to its biodiversity. Climate change represents a major challenge to the survival and viability of species, and the potential consequences must be assessed to allow for mitigation. We project the effects of several climate change scenarios on bat diversity, and predict changes in range size for 171 bat species throughout SEA. We predict decreases in species richness in all areas with high species richness (>80 species) at 2050–2080, using bioclimatic IPCC scenarios A2 (a severe scenario, continuously increasing human population size, regional changes in economic growth) and B1 (the ‘greenest’ scenario, global population peaking mid‐century). We also predicted changes in species richness in scenarios that project vegetation changes in addition to climate change up to 2050. At 2050 and 2080, A2 and B1 scenarios incorporating changes in climatic factors predicted that 3–9% species would lose all currently suitable niche space. When considering total extents of species distribution in SEA (including possible range expansions), 2–6% of species may have no suitable niche space in 2050–2080. When potential vegetation and climate changes were combined only 1% of species showed no changes in their predicted ranges by 2050. Although some species are projected to expand ranges, this may be ecologically impossible due to potential barriers to dispersal, especially for species with poor dispersal ability. Only 1–13% of species showed no projected reductions in their current range under bioclimatic scenarios. An effective way to facilitate range shift for dispersal‐limited species is to improve landscape connectivity. If current trends in environmental change continue and species cannot expand their ranges into new areas, then the majority of bat species in SEA may show decreases in range size and increased extinction risk within the next century.     

Effects of simulated long-term climatic change on the bryophytes of a limestone grassland community

The bryophyte vegetation of upland limestone grassland at Buxton in the southern Pennine Hills (UK) was studied following seven years' continuous simulated climate change treatments. The experimental design involved two temperature regimes (ambient, winter warming by 3°C) in factorial combination with three moisture regimes (normal, summer drought, supplemented summer rainfall) and with five replicate blocks. Percentage cover of the bryophytes was estimated visually using 15 randomly positioned quadrats (30 cm × 30 cm) within each of the 30 3 m × 3 m plots. Significant treatment effects were found but these were relatively modest. Total bryophyte cover and cover of Calliergonella cuspidata and Rhytidiadelphus squarrosus responded negatively to drought, whereas Fissidens dubius increased in the droughted plots. Campyliadelphus chrysophyllus increased with winter warming, while R. squarrosus, Lophocolea bidentata and species richness all decreased. The effects on the total bryophyte flora were further studied by canonical correspondence analysis, which yielded a first axis reflecting the combined effects of the moisture and temperature treatments. However, this analysis and a detrended correspondence analysis of the plot data also revealed that natural factors were more important causes of variation in the grassland community than the simulated climate treatments. It was concluded that dewfall may be an important source of moisture for grassland bryophytes and that this factor may have reduced the impact of the moisture treatments. The absence of some thermophilous species such as Homalothecium lutescens in the plots initially may also have reduced their scope for major vegetational change.     

Phenotype microarray profiling of Staphylococcus aureus menD and hemB mutants with the small-colony-variant phenotype

Standard biochemical tests have revealed that hemin and menadione auxotrophic Staphylococcus aureus small-colony variants (SCVs) exhibit multiple phenotypic changes. To provide a more complete analysis of the SCV phenotype, two genetically defined mutants with a stable SCV phenotype were comprehensively tested. These mutants, generated via mutations in menD or hemB that yielded menadione and hemin auxotrophs, were subjected to phenotype microarray (PM) analysis of over 1,500 phenotypes (including utilization of different carbon, nitrogen, phosphate, and sulfur sources; growth stimulation or inhibition by amino acids and other nutrients, osmolytes, and metabolic inhibitors; and susceptibility to antibiotics). Compared to parent strain COL, the hemB mutant was defective in utilization of a variety of carbon sources, including Krebs cycle intermediates and compounds that ultimately generate ATP via electron transport. The phenotype of the menD mutant was similar to that of the hemB mutant, but the defects in carbon metabolism were more pronounced than those seen with the hemB mutant. In both mutant strains, hexose phosphates and other carbohydrates that provide ATP in the absence of electron transport stimulated growth. Other phenotypes of SCV mutants, such as hypersensitivity to sodium selenite, sodium tellurite, and sodium nitrite, were also uncovered by the PM analysis. Key results of the PM analysis were confirmed in independent growth studies and by using Etest strips for susceptibility testing. PM technology is a new and efficient technology for assessing cellular phenotypes in S. aureus.     

Preparation, stability, and in vitro performance of vesicles made with diblock copolymers

Vesicles made completely from diblock copolymers-polymersomes-can be stably prepared by a wide range of techniques common to liposomes. Processes such as film rehydration, sonication, and extrusion can generate many-micron giants as well as monodisperse, approximately 100 nm vesicles of PEO-PEE (polyethyleneoxide-polyethylethylene) or PEO-PBD (polyethyleneoxide-polybutadiene). These thick-walled vesicles of polymer can encapsulate macromolecules just as liposomes can but, unlike many pure liposome systems, these polymersomes exhibit no in-surface thermal transitions and a subpopulation even survive autoclaving. Suspension in blood plasma has no immediate ill-effect on vesicle stability, and neither adhesion nor stimulation of phagocytes are apparent when giant polymersomes are held in direct, protracted contact. Proliferating cells, in addition, are unaffected when cultured for an extended time with an excess of polymersomes. The effects are consistent with the steric stabilization that PEG-lipid can impart to liposomes, but the present single-component polymersomes are far more stable mechanically and are not limited by PEG-driven micellization. The results potentiate a broad new class of technologically useful, polymer-based vesicles.