Solution structure of the Escherichia coli protein ydhR: a putative mono-oxygenase

YdhR is a 101-residue conserved protein from Escherichia coli. Sequence searches reveal that the protein has >50% identity to proteins found in a variety of other bacterial genomes. Using size exclusion chromatography and fluorescence spectroscopy, we determined that ydhR exists in a dimeric state with a dissociation constant of approximately 40 nM. The three-dimensional structure of dimeric ydhR was determined using NMR spectroscopy. A total of 3400 unambiguous NOEs, both manually and automatically assigned, were used for the structure calculation that was refined using an explicit hydration shell. A family of 20 structures was obtained with a backbone RMSD of 0.48 A for elements of secondary structure. The structure reveals a dimeric alpha,beta fold characteristic of the alpha+beta barrel superfamily of proteins. Bioinformatic approaches were used to show that ydhR likely belongs to a recently identified group of mono-oxygenase proteins that includes ActVA-Orf6 and YgiN and are involved in the oxygenation of polyaromatic ring compounds.     

High-resolution aliphatic side-chain assignments in 3D HCcoNH experiments with joint H–C evolution and non-uniform sampling

We describe an efficient NMR triple resonance approach that correlates, at high resolution, protein side-chain and backbone resonances. It relies on the combination of two strategies: joint evolution of aliphatic side-chain proton/carbon coherences using a backbone N–H based HCcoNH reduced dimensionality (RD) experiment and non-uniform sampling (NUS) in two indirect dimensions. A typical data set containing such correlation information can be acquired in 2 days, at very high resolution unfeasible for conventional 4D HCcoNH-TOCSY experiments. The resonances of the aliphatic side-chain protons are unambiguously assigned to their attached carbons through the analysis of the ‘sum’ and ‘difference’ spectra. This approach circumvents the tedious process of manual resonance assignments using HCcH-TOCSY data, while providing additional resolving power of backbone N–H signals. A simple peak-list based algorithm has been implemented in the IBIS software for rapid automated backbone and side-chain assignments.     

Earliest record of Platychoerops (Primates,Plesiadapidae), a new species from Mouras Quarry,Mont de Berru,France

Plesiadapids are extinct relatives of extant euarchontans (primates, dermopterans, and scandentians), which lived in North America and Europe during the Paleocene and Early Eocene. The only genus of plesiadapid whose species are absent from Paleocene strata is Platychoerops. Further, Platychoerops is the only group found in sediments post‐dating the Paleocene‐Eocene boundary (PEB) by a substantial period of time based on large samples. It is also substantially different from other plesiadapids in dental features thought to reflect ecology. Its evolution has been linked to the rapid global climate change and faunal turnover marking the PEB. Platychoerops and Plesiadapis tricuspidens have been reconstructed as members of a single lineage by some authors. We describe a specimen (right p3‐m3) that we attribute to a new species, Platychoeropsantiquus, from the unequivocally Paleocene (MP6) Mouras Quarry of Mont de Berru, France. It has strong morphological affinities to Platychoerops daubrei yet co‐occurs with many specimens of Plesiadapis tricuspidens, as well as the plesiadapid Chiromyoides campanicus. The existence of a species of Platychoerops prior to the PEB decouples the evolution of Platychoerops from the climate change and faunal turnover event associated with the PEB. Furthermore, the co‐occurrence of Platychoerops with P. tricuspidens refutes the idea of a single lineage for these taxa. Instead, Platychoerops may be more closely related to North American Plesiadapis cookei (a previous alternate hypothesis). We suggest character displacement in a Paleocene immigrant population of P. cookei resulting from competition with sympatric P. tricuspidens, as a likely scenario for the evolution of Platychoerops. Am J Phys Anthropol, 2012. © 2012 Wiley Periodicals, Inc.     

Detection of HbA1c by boronate affinity immunoassay using bacterial magnetic particles

We have developed a boronate affinity immunoassay system using m-aminophenylboronic acid (mAPB) coupling to bacterial magnetic particles (BMPs). Homobifunctional crosslinker, Bis-(succcimidyl)suberate (BS3), was employed for preparation of mAPB-BMPs conjugates (mAPB-BMPs). Quantities of HbA_1c on mAPB-BMPs were evaluated based on luminescence from alkaline phosphatase-conjugated anti-Hb antibody (ALP–antibody) binding to HbA_1c on the BMP surface. The binding of HbA_1c to mAPB-BMPs occurred gradually and was almost completed within 10 mm. The coupling reaction is enhanced due to static electric interaction between the positive charges on HbA_1c and negative charges on BMPs. The amount of HbA_1c binding to mAPB-BMPs increased with increasing sodium chloride concentrations in the range of 0–100 mM. However, the amount of Hb binding to mAPB-BMPs also increased in high concentration of sodium chloride. The Hb binding to mAPB-BMPs was detached from mAPB-BMPs when Hb–mAPB-BMPs were washed with low salt buffer. This indicates that Hb is nonspecifically adsorbed onto the surface of mAPB-BMPs in high concentration of sodium chloride. These results suggest that selective separation of HbA_1c using mAPB-BMPs can be achieved with these conditions. A dose–response curve was obtained between luminescence intensity and HbA_1c concentration using a fully automated boronate affinity immunoassay. A linear relationship between luminescence intensity and HbA_1c concentration was obtained in the range of 10–10⁴ ng/ml.     

Early queen infection shapes developmental dynamics and induces long-term disease protection in incipient ant colonies

Infections early in life can have enduring effects on an organism's development and immunity. In this study, we show that this equally applies to developing ‘superorganisms’––incipient social insect colonies. When we exposed newly mated Lasius niger ant queens to a low pathogen dose, their colonies grew more slowly than controls before winter, but reached similar sizes afterwards. Independent of exposure, queen hibernation survival improved when the ratio of pupae to workers was small. Queens that reared fewer pupae before worker emergence exhibited lower pathogen levels, indicating that high brood rearing efforts interfere with the ability of the queen's immune system to suppress pathogen proliferation. Early-life queen pathogen exposure also improved the immunocompetence of her worker offspring, as demonstrated by challenging the workers to the same pathogen a year later. Transgenerational transfer of the queen's pathogen experience to her workforce can hence durably reduce the disease susceptibility of the whole superorganism.     

Predictable shorebird departure patterns from a staging site can inform collision risks and mitigation of wind energy developments

High-quality staging sites are critical for long-distance migratory shorebirds to rest and refuel but are under threat from human development, including expansion of wind energy projects. However, predicting migration timing and movements in relation to weather conditions at staging sites can increase our understanding and mitigate effects of wind turbine collisions. Here we assessed northward migration timing and orientation in relation to environmental conditions at an inland staging area in Saskatchewan, Canada, with active and proposed wind energy developments. The area is known to host ~25% of North America's Sanderling Calidris alba population and 16 other Arctic-breeding migrant shorebird species. We quantified arrival and departure time of day in relation to weather using data from 140 of 237 Sanderlings radiotagged locally and at a southern staging site in the Gulf of Mexico with the Motus Wildlife Tracking System (April–June, 2015–2017). Although Sanderling arrival times were not related to time of day or weather, departures were more likely at sunset in winds blowing towards the northwest at intermediate speeds (<22 km/h). Departure flights were also primarily oriented north-northwest in the direction of a proposed wind energy development site at a mean ground speed of 21.4 m/s. Based on published climb rates and flight speed data, we estimated that shorebirds needed between 2 and 14 km setback distance to clear maximum turbine heights of 165 m. Given that departure events were predictable in time and space, adaptive mitigation may be useful for planning wind energy developments while reducing risk for staging Arctic-breeding shorebirds.     

Influence of the completeness of chemical shift assignments on NMR structures obtained with automated NOE assignment

Reliable automated NOE assignment and structure calculation on the basis of a largely complete, assigned input chemical shift list and a list of unassigned NOESY cross peaks has recently become feasible for routine NMR protein structure calculation and has been shown to yield results that are equivalent to those of the conventional, manual approach. However, these algorithms rely on the availability of a virtually complete list of the chemical shifts. This paper investigates the influence of incomplete chemical shift assignments on the reliability of NMR structures obtained with automated NOESY cross peak assignment. The program CYANA was used for combined automated NOESY assignment with the CANDID algorithm and structure calculations with torsion angle dynamics at various degrees of completeness of the chemical shift assignment which was simulated by random omission of entries in the experimental ¹H chemical shift lists that had been used for the earlier, conventional structure determinations of two proteins. Sets of structure calculations were performed choosing the omitted chemical shifts randomly among all assigned hydrogen atoms, or among aromatic hydrogen atoms. For comparison, automated NOESY assignment and structure calculations were performed with the complete experimental chemical shift but under random omission of NOESY cross peaks. When heteronuclear-resolved three-dimensional NOESY spectra are available the current CANDID algorithm yields in the absence of up to about 10% of the experimental ¹H chemical shifts reliable NOE assignments and three-dimensional structures that deviate by less than 2 Å from the reference structure obtained using all experimental chemical shift assignments. In contrast, the algorithm can accommodate the omission of up to 50% of the cross peaks in heteronuclear- resolved NOESY spectra without producing structures with a RMSD of more than 2 Å to the reference structure. When only homonuclear NOESY spectra are available, the algorithm is slightly more susceptible to missing data and can tolerate the absence of up to about 7% of the experimental ¹H chemical shifts or of up to 30% of the NOESY peaks.Abbreviations: BmPBP^A – Bombyx mori pheromone binding protein form A; CYANA – combined assignment and dynamics algorithm for NMR applications; NMR – nuclear magnetic resonance; NOE – nuclear Overhauser effect; NOESY – NOE spectroscopy; RMSD – root-mean-square deviation; WmKT – Williopsis mrakii killer toxin