Ladderane phospholipids in anammox bacteria comprise phosphocholine and phosphoethanolamine headgroups

Anammox bacteria present in wastewater treatment systems and marine environments are capable of anaerobically oxidizing ammonium to dinitrogen gas. This anammox metabolism takes place in the anammoxosome which membrane is composed of lipids with peculiar staircase-like 'ladderane' hydrocarbon chains that comprise three or four linearly concatenated cyclobutane structures. Here, we applied high-performance liquid chromatography coupled to electrospray ionization tandem mass spectrometry and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry to elucidate the full identity of these ladderane lipids. This revealed a wide variety of ladderane lipid species with either a phosphocholine or phosphoethanolamine polar headgroup attached to the glycerol backbone. In addition, in silico analysis of genome data gained insight into the machinery for the biosynthesis of the phosphocholine and phosphoethanolamine phospholipids in anammox bacteria.     

Exploring the recognition of quadruplex DNA by an engineered Cys2-His2 zinc finger protein

We have recently described an engineered zinc finger protein (Gq1) that binds with high specificity to the intramolecular G-quadruplex formed by the human telomeric sequence 5'-(GGTTAG)(5)-3', and that inhibits the activity of the enzyme telomerase in vitro. Here we report site-directed mutagenesis, biophysical, and molecular modeling studies that provide new insights into quadruplex recognition by the zinc finger scaffold. We show that any one finger of Gq1 can be replaced with the corresponding finger of Zif268, without significant loss of quadruplex affinity or quadruplex versus duplex discrimination. Replacement of two fingers, with one being finger 2, of Gq1 by Zif268 results in significant impairment of quadruplex recognition and loss of discrimination. Molecular modeling suggests that the zinc fingers of Gq1 can bind to the human parallel-stranded quadruplex structure in a stable arrangement, whereas Zif268-quadruplex models show significantly weaker binding energy. Modeling also suggests that an important role of the key protein finger residues in the Gq1-quadruplex complex is to maintain Gq1 in an optimum conformation for quadruplex recognition.     

‘Bioenergy from cattle manure? Implications of anaerobic digestion and subsequent pyrolysis for carbon and nitrogen dynamics in soil’

Cattle manure can be processed to produce bioenergy, resulting in by‐products with different physicochemical characteristics. To evaluate whether application of such bioenergy by‐products to soils would be beneficial compared with their unprocessed counterpart, we quantified differences in greenhouse gas emissions and carbon (C) and nitrogen (N) dynamics in soil. Three by‐products (¹⁵N‐labeled cattle manure, from which anaerobic digestate was obtained, which was subsequently pyrolysed) were applied to a loess and a sandy soil in a laboratory incubation study. The highest losses of soil C from biological activity (CO₂ respiration) were observed in manure treatments (39% and 32% for loess and sandy soil), followed by digestate (31% and and 18%), and biochar (15% and and 7%). Emissions of nitrous oxide (N₂O) ranged from 0.6% of applied N from biochar to 4.0% from manure. Isotope labeling indicated that manure N was most readily mineralized, contributing 50% to soil inorganic N. The anaerobic digestate was the only by‐product increasing the mineral N pool, while reducing emissions of N₂O compared with manure. In biochar treatments, less than 18.3% of soil mineral N derived from the biochar, while it did not constrain mineralization of native soil N. By‐products of anaerobic digestion and pyrolysis revealed soil fertility in addition to environmental benefits. However, the reported advantages lessen when the declining yields of C and N over the bioenergy chain are considered.     

Are you approaching me? Motor execution influences perceived action orientation

Human observers are especially sensitive to the actions of conspecifics that match their own actions. This has been proposed to be critical for social interaction, providing the basis for empathy and joint action. However, the precise relation between observed and executed actions is still poorly understood. Do ongoing actions change the way observers perceive others' actions? To pursue this question, we exploited the bistability of depth-ambiguous point-light walkers, which can be perceived as facing towards the viewer or as facing away from the viewer. We demonstrate that point-light walkers are perceived more often as facing the viewer when the observer is walking on a treadmill compared to when the observer is performing an action that does not match the observed behavior (e.g., cycling). These findings suggest that motor processes influence the perceived orientation of observed actions: Acting observers tend to perceive similar actions by conspecifics as oriented towards themselves. We discuss these results in light of the possible mechanisms subtending action-induced modulation of perception.     

Spinal reflex properties in the long term after stroke

In the long term after stroke, secondary functional deterioration may be observed while patients also get older. Possible underlying mechanisms are largely unknown. We aimed to assess neuromuscular degeneration represented by alterations in peripheral reflex loop characteristics as a function of follow-up time after stroke, controlled for age. Twenty-one stroke survivors within a small age range (62-67 years) but large variance in follow-up time after stroke (1-15 years) and both five age matched (59-62 years) and young subjects (28-36 years) participated. Short and long latency reflexes evoked by ramp and hold stretches were identified from EMG traces of the m. flexor carpi radialis. Short latency reflex onset time was not enhanced (mean difference 1.6ms compared to age matched controls) and did not relate to follow-up time after stroke (p=0.81). Young controls showed significantly lower reflex delay times (mean difference 7.2ms with respect to older subjects, p=0.009). No evidence was found for peripheral neuromuscular deterioration as a function of follow up time after stroke. Functional deterioration as a result of ageing of stroke patients that may interact with post stroke follow-up time is of further interest.     

Kinetic characterization of lipid II-Ala:alanyl-tRNA ligase (MurN) from Streptococcus pneumoniae using semisynthetic aminoacyl-lipid II substrates

MurM and MurN are tRNA-dependent ligases that catalyze the addition of the first (L-Ala/L-Ser) and second (L-Ala) amino acid onto lipid II substrates in the biosynthesis of the peptidoglycan layer of Streptococcus pneumoniae. We have previously characterized the first ligase, MurM (Lloyd, A. J., Gilbey, A. M., Blewett, A. M., De Pascale, G., El Zoeiby, A., Levesque, R. C., Catherwood, A. C., Tomasz, A., Bugg, T. D., Roper, D. I., and Dowson, C. G. (2008) J. Biol. Chem. 283, 6402-6417). In order to characterize the second ligase MurN, we have developed a chemoenzymatic route to prepare the lipid II-Ala and lipid II-Ser substrates. Recombinant MurN enzymes from penicillin-resistant (159) and -sensitive (Pn16) S. pneumoniae were expressed and purified as MBP fusion proteins and reconstituted using a radiochemical assay. MurN ligases from strains 159 and Pn16 both showed a 20-fold higher catalytic efficiency for lipid II-L-Ala over lipid II-l-Ser, with no activity against unmodified lipid II, and similar kinetic parameters were measured for MurN from penicillin-resistant and penicillin-sensitive strains. These results concur with the peptidoglycan analysis of S. pneumoniae, in which the major cross-link observed is L-Ala-L-Ala. The combined action of ligases MurM and MurN is therefore required in order to rationalize the high level of dipeptide cross-links in penicillin-resistant S. pneumoniae, with ligase MurM showing the major difference between penicillin-resistant and penicillin-sensitive strains.