Ternary structure reveals mechanism of a membrane diacylglycerol kinase

Diacylglycerol kinase catalyses the ATP-dependent conversion of diacylglycerol to phosphatidic acid in the plasma membrane of Escherichia coli. The small size of this integral membrane trimer, which has 121 residues per subunit, means that available protein must be used economically to craft three catalytic and substrate-binding sites centred about the membrane/cytosol interface. How nature has accomplished this extraordinary feat is revealed here in a crystal structure of the kinase captured as a ternary complex with bound lipid substrate and an ATP analogue. Residues, identified as essential for activity by mutagenesis, decorate the active site and are rationalized by the ternary structure. The g-phosphate of the ATP analogue is positioned for direct transfer to the primary hydroxyl of the lipid whose acyl chain is in the membrane. A catalytic mechanism for this unique enzyme is proposed. The active site architecture shows clear evidence of having arisen by convergen

The bacterial stressosome:a modular system that has been adapted to control secondary messenger signaling

SummaryThe stressosome complex regulates downstream effectors in response to environmental signals. In Bacillus subtilis, it activates the alternative sigma factor σB, leading to the upregulation of the general stress regulon. Herein, we characterize a stressosome-regulated biochemical pathway in Moorella thermoacetica. We show that the presumed sensor, MtR, and the scaffold, MtS, form a pseudo-icosahedral structure like that observed in B. subtilis. The N-terminal domain of MtR is structurally homologous to B. subtilis RsbR, despite low sequence identity. The affinity of the switch kinase, MtT, for MtS decreases following MtS phosphorylation and not because of structural reorganization. Dephosphorylation of MtS by the PP2C type phosphatase MtX permits the switch kinase to rebind the stressosome to reset the response. We also show that MtT regulates cyclic di-GMP biosynthesis through inhibition of a GG(D/E)EF-type diguanylate cyclase, demonstrating that secondary messenger levels are regulated by the stressosome

Genomic Insights Into The Ixodes scapularis Tick Vector Of Lyme Disease

Ticks transmit more pathogens to humans and animals than any other arthropod. We describe the 2.1 Gbp nuclear genome of the tick, Ixodes scapularis (Say), which vectors pathogens that cause Lyme disease, human granulocytic anaplasmosis, babesiosis and other diseases. The large genome reflects accumulation of repetitive DNA, new lineages of retrotransposons, and gene architecture patterns resembling ancient metazoans rather than pancrustaceans. Annotation of scaffolds representing B57% of the genome, reveals 20,486 protein-coding genes and expansions of gene families associated with tick–host interactions. We report insights from genome analyses into parasitic processes unique to ticks, including host ‘questing’, prolonged feeding, cuticle synthesis, blood meal concentration, novel methods of haemoglobin digestion, haem detoxification, vitellogenesis and prolonged off-host survival. We identify proteins associated with the agent of human granulocytic anaplasmosis, an emerging disease, and the encephalitis-causing Langat virus, and a population structure correlated to life-history traits and transmission of the Lyme disease agent

Genomic Insights Into The Ixodes scapularis Tick Vector Of Lyme Disease

Ticks transmit more pathogens to humans and animals than any other arthropod. We describe the 2.1 Gbp nuclear genome of the tick, Ixodes scapularis (Say), which vectors pathogens that cause Lyme disease, human granulocytic anaplasmosis, babesiosis and other diseases. The large genome reflects accumulation of repetitive DNA, new lineages of retrotransposons, and gene architecture patterns resembling ancient metazoans rather than pancrustaceans. Annotation of scaffolds representing B57% of the genome, reveals 20,486 protein-coding genes and expansions of gene families associated with tick–host interactions. We report insights from genome analyses into parasitic processes unique to ticks, including host ‘questing’, prolonged feeding, cuticle synthesis, blood meal concentration, novel methods of haemoglobin digestion, haem detoxification, vitellogenesis and prolonged off-host survival. We identify proteins associated with the agent of human granulocytic anaplasmosis, an emerging disease, and the encephalitis-causing Langat virus, and a population structure correlated to life-history traits and transmission of the Lyme disease agent

Assessment of protein-protein interfaces in cryo-EM derived assemblies

Structures of macromolecular assemblies derived from cryo-EM maps often contain errors that become more abundant with decreasing resolution. Despite efforts in the cryo-EM community to develop metrics for map and atomistic model validation, thus far, no specific scoring metrics have been applied systematically to assess the interface between the assembly subunits. Here, we comprehensively assessed protein–protein interfaces in macromolecular assemblies derived by cryo-EM. To this end, we developed Protein Interface-score (PI-score), a density-independent machine learning-based metric, trained using the features of protein–protein interfaces in crystal structures. We evaluated 5873 interfaces in 1053 PDB-deposited cryo-EM models (including SARS-CoV-2 complexes), as well as the models submitted to CASP13 cryo-EM targets and the EM model challenge. We further inspected the interfaces associated with low-scores and found that some of those, especially in intermediate-to-low resolution (worse than 4 Å) structures, were not captured by density-based assessment scores. A combined score incorporating PI-score and fit-to-density score showed discriminatory power, allowing our method to provide a powerful complementary assessment tool for the ever-increasing number of complexes solved by cryo-EM

Plant management as an integrated part of Ireland's aquatic resources

Aquatic plants are fundamental to the structure and functioning of many freshwater habitats. In undisturbed aquatic habitats plants serve a multitude of important functions including the provision of substrates for colonisation by periphyton and macro-invertebrates, a direct and indirect food source for macro-invertebrates, fish and wildfowl, spawning substrates for coarse fish species, shelter for young and adult fish oxygen as a by-product of photosynthesis. In disturbed habitats, however, the natural balance among aquatic plants is altered and an excessive, often monospecific, vegetation commonly develops. This can adversely affect the beneficial use of the watercourse by diminishing its amenity and recreational potential, exacerbating water supply and flooding problems and resulting in excessive siltation in river or canal habitats. This paper outlines the nature and extent of the more important weed problems in lrish aquatic situations and describes a range of control strategies that may be adopted to rehabilitate these habitats and to enable their exploitation by a diversity of user groups. In certain instances paucity of aquatic vegetation presents problems, including bankside destabilisation and inchannel siltation. Reed transplantation trials have been conducted in denuded lrish canals in an effort to rehabilitate these habitats and the results from these are presented. Weed control procedures generally embrace four broad categories. These are mechanical, chemical, environmental and biological control. Of these only biological control is not widely adopted in Ireland. The effects that these procedures have on specific weeds were quantitatively investigated at a number of aquatic situations throughout the country. Results from experiments among a diversity of aquatic plant species and involving the use of mechanical weed cutting apparatus show that, unless the plant is cut to a depth at which light penetration is significantly diminished, rapid regrowth will occur and higher standing crops than might otherwise be expected may be recorded. Experimentation with a small number of herbicides has revealed that, when used with discrimination, these provide effective weed control, with little or no detrimental effect to the aquatic ecosystem. Results from trials with dichlobenil and glyphosate on aquatic and riparian species are presented. The effect that shading has on macrophyte biomass and the effect that rotted barley straw has on algal growths in watercourses is also described

Screening of acyl hydrazide proteinase inhibitors for antiparasitic activity against Trypanosoma brucei

The major cysteine proteinase (brucipain) of Trypanosoma brucei is a target for chemotherapy of African Sleeping Sickness. We have screened a non-peptidyl acyl hydrazide proteinase inhibitor library of 500 compounds for inhibition of brucipain. Those 21 compounds with IC(50) values of <40 microM were tested for efficacy against bloodstream forms of T. brucei in cell culture. Eight acyl hydrazides showed 50% or more inhibition of trypanosome replication at <1 microM. The trypanocidal acitivity of the most effective compounds was comparable with those of the commercial antitrypanosomal drugs suramin and diminazene aceturate. However, these acyl hydrazides exhibited varying cytotoxicity towards human HL-60 cells and therefore, only less favourable selectivity indices compared with the commercially available drugs. Nevertheless, the data support the potential of acyl hydrazides as antitrypanosomal chemotherapeutic agents for treatment of sleeping sickness