Caffeine Junkie: an Unprecedented Glutathione S-Transferase-Dependent Oxygenase Required for Caffeine Degradation by Pseudomonas putida CBB5

Caffeine and other N-methylated xanthines are natural products found in many foods, beverages, and pharmaceuticals. Therefore, it is not surprising that bacteria have evolved to live on caffeine as a sole carbon and nitrogen source. The caffeine degradation pathway of Pseudomonas putida CBB5 utilizes an unprecedented glutathione-S-transferase-dependent Rieske oxygenase for demethylation of 7-methylxanthine to xanthine, the final step in caffeine N-demethylation. The gene coding this function is unusual, in that the iron-sulfur and non-heme iron domains that compose the normally functional Rieske oxygenase (RO) are encoded by separate proteins. The non-heme iron domain is located in the monooxygenase, ndmC, while the Rieske [2Fe-2S] domain is fused to the RO reductase gene, ndmD. This fusion, however, does not interfere with the interaction of the reductase with N₁- and N₃-demethylase RO oxygenases, which are involved in the initial reactions of caffeine degradation. We demonstrate that the N₇-demethylation reaction absolutely requires a unique, tightly bound protein complex composed of NdmC, NdmD, and NdmE, a novel glutathione-S-transferase (GST). NdmE is proposed to function as a noncatalytic subunit that serves a structural role in the complexation of the oxygenase (NdmC) and Rieske domains (NdmD). Genome analyses found this gene organization of a split RO and GST gene cluster to occur more broadly, implying a larger function for RO-GST protein partners.     

Mutation K42R in Ribosomal Protein S12 Does Not Affect Susceptibility of Mycobacterium smegmatis 16S rRNA A-Site Mutants to 2-Deoxystreptamines

Recent studies have suggested that ribosomal protein S12 modulates 16S rRNA function and susceptibility to 2-deoxystreptamine aminoglycosides. To study whether the non-restrictive K42R mutation in RpsL affects 2-deoxystreptamine susceptibility in Mycobacterium smegmatis, we studied the drug susceptibility pattern of various mutants with genetic alterations in the 16S rRNA decoding A-site in the context of wild-type and mutant protein S12. RpsL K42R substitution was found not to affect the drug resistance pattern associated with mutational alterations in 16S rRNA H44.     

Failure of duplexes based on polylaurusin [poly(L), “polyformycin B”] to induce interferon

Polylaurusin[poly(L) or “polyformycin B”] forms double-stranded complexes with polycytidylic acid (poly(C)) and with poly(5-bromocytidylic acid) [poly(br⁵C)] with T_m's of 46.5° (0.2 $\text{M}$ NaCl, pH 7) and 72.5° (0.15 M NaCl, pH 7), respectively. Both complexes fail to provide antiviral resistance (against vesicular stomatitis virus in primary rabbit kidney cells) or to induce interferon in “superinduced” primary rabbit kidney cells, even though they fulfill all previously recognized requirements for effective interferon inducers.     

Structure of the stridulation ofOrchelimum (Orthoptera: Tettigoniidae)

Summary The structure of the stridulation was investigated by re-playing tape-recordings at very slow speed. The findings were corrobrated by sonograms and mingograms.The central part of the song is the ripple, a fast succession of syllables around which isolated syllables (clicks) are distributed according to species and circumstances. The rate of syllables in the ripple is a linear function of temperature.A quantitative expression for the stridulatory activity is the actual number of syllables per time unit, including pauses. By changes in the combination of elements, at leastO. agile is able to increase the output of syllables four to six times.This work was made possible by a grant from the Carlsberg Foundation to whom my most sincere thanks are due. —As I do not have advanced equipment for sound analysis, I am very much indebted to Dr. Bondesen and cand. sci. Poul Hansen, Bioakustisk Laboratorium, Naturhistorisk Museum, Aarhus, Denmark, and Mr. W.B. Broughton and Dr. M. Samways, Animal Acoustic Unit, City of London Polytechnic, London, for analysing part of the material by sonograms and mingograms. The very valuable help of Dr. Th.J. Walker, University of Florida, Gainesville, Florida, in identifying the species is gratefully acknowledged. For friendly discussions and linguistic corrections my best thanks are due to H.T. Evans and F.D.S. Evans.     

Diversity,dynamics and reproduction in a community of small mammals in Upper Guinea,with emphasis on pygmy mice ecology

As part of a large survey on reservoirs of Lassa fever in Guinea, three villages were investigated in high endemic zone, close to Sierra Leone border. Biodiversity of the small mammal community is presented in this study through a standardized trapping in houses, cultivations and forest. Identification of the small mammals was based on morphology and by molecular technique for sibling species. Of the 1123 specimens collected in 2003–2005, we identified seventeen species (thirteen Muridae, four Soricidae), leading to high diversity (Shannon index = 1.6–1.8) and high equitability (evenness index = 0.7–0.8) in cultivations and forest. In houses conversely, the rodent community was dominated by Mastomys natalensis (95–98%), leading to low diversity and equitability. Dynamics and reproduction were investigated in two species of pygmy mice, Mus mattheyi and Mus minutoides, two species of Praomys, P. daltoni and P. rostratus, and in Mastomys erythroleucus. The pygmy mice were abundant in cultivations in early rainy season, and reproduced from rainy to dry season. Praomys daltoni was also found more abundant in cultivations and seemed to reproduce between rainy and dry season, whereas P. rostratus preferred forest and cultivations in late rainy season, and reproduced throughout the year. Finally, M. erythroleucus was more abundant in forest in dry season, and seemed to reproduce from late rainy to dry season. This species had a low occurrence (6.5%) in the Faranah’s zone, and probably lived at its southern limit in Guinea. The presence of other Murinae, such as M. natalensis, Praomys spp as possible competitors in the same habitats, is discussed. For the first time, this study relates population biology of pygmy mice with molecular identification.     

The Application of Electron Backscatter Diffraction on Halide Perovskite Materials

Recently, the application of electron backscatter diffraction (EBSD) in halide perovskites has enabled the correlation of the micro‐structural arrangement of polycrystalline grains with other properties (optical, electrical, mechanical, and chemical) in a “pixel‐by‐pixel” approach. Most studies so far have used an ultra‐sensitive electron beam detector that has sensitivity thousands of times higher than a traditional scintillator screen and charge coupled device camera, enabling much lower beam currents. An alternative approach has been the use of low vacuum measurement conditions to avoid charge buildup that leads to damage. This review focuses on introducing the classical EBSD technique to the halide perovskite community, where it has been highly underutilized due to beaminduced damage in these relatively unstable materials. Recent research is used to dispel some common misconceptions about grain boundaries in halide perovskites and highlight what has been learned by comparing and correlating EBSD with other techniques. Additionally, the remaining limitations, development challenges, and future of the EBSD technique for halide perovskites are discussed. Successful utilization of the EBSD technique as a common characterization tool in the halide perovskite community will enable scientists and engineers to develop maps of cross correlated properties, helping to unlock the full potential of this complex material system.     

Synthesis of homoagmatine and GC–MS analysis of tissue homoagmatine and agmatine: evidence that homoagmatine but not agmatine is a metabolite of pharmacological L-homoarginine in the anesthetized rat

Amino Acids - Low L-homoarginine (hArg) concentrations in human blood and urine are associated with renal and cardiovascular morbidity and mortality, yet the underlying mechanisms and the...     

Habitat fragmentation shapes natal dispersal and sociality in an Afrotropical cooperative breeder

It remains poorly understood how effects of anthropogenic activity, such as large-scale habitat fragmentation, impact sociality in animals. In cooperatively breeding species, groups are mostly formed through delayed offspring dispersal, and habitat fragmentation can affect this process in two opposite directions. Increased habitat isolation may increase dispersal costs, promoting delayed dispersal. Alternatively, reduced patch size and quality may decrease benefits of philopatry, promoting dispersal. Here, we test both predictions in a cooperatively breeding bird (placid greenbul, Phyllastrephus placidus) from an Afrotropical cloud forest archipelago. Males born in fragmented forest dispersed about 1 year earlier than those born in continuous forest. Contrary to females, males also started to reproduce earlier and mostly settled within their natal patch. Females only rarely delayed their dispersal for more than 1 year, both in fragmented and continuous forests. Our results suggest that early male dispersal and reproduction is jointly driven by a decrease in the value of the natal territory and an increase in local breeding opportunities in fragmented forest. While plasticity in dispersal strategies of cooperative breeders in response to anthropogenic change is believed to optimize reproduction-survival trade-offs, to what extent it shapes the ability of species to respond to rapid environmental change remains to be studied.     

Modulation of transmembrane pressure in manufacturing scale tangential flow filtration N-1 perfusion seed culture

Mammalian cells were grown to high density in a 3,000 L culture using perfusion with hollow fibers operated in a tangential flow filtration mode. The high-density culture was used to inoculate the production stage of a biomanufacturing process. At constant permeate flux operation, increased transmembrane pressures (TMPs) were observed on the final day of the manufacturing batches. Small scale studies suggested that the filters were not irreversibly fouled, but rather exposed to membrane concentration polarization that could be relieved by tangential sweeping of the hollow fibers. Studies were undertaken to analyze parameters that influence the hydrodynamic profile within hollow fibers; including filter area, cell density, recirculation flow rate, and permeate flow rate. Results indicated that permeate flow rate had the greatest influence on modulating TMP. Further evaluation showed a significant decrease in TMP when permeate flow was reduced, and this occurred without any negative effect on cell growth or viability. Hence, a 30% reduction of permeate flow rate was implemented at manufacturing scale. A stable operation was achieved as TMP was successfully reduced by 75% while preserving all critical factors for performance in the perfusion bioreactor.