Secondary structure and NMR resonance assignments of the C-terminal DNA-binding domain of Uup protein

ATP-binding cassette (ABC) systems belong to a large superfamily of proteins that couple the energy released from ATP hydrolysis to a wide variety of cellular processes, including not only transport of various molecules, but also gene regulation, and DNA repair. Mutations in the bacterial uup gene, which encodes a cytosolic ABC ATPase, lead to an increase in the frequency of precise excision of transposons Tn10 and Tn5, suggesting a role of the Uup protein in DNA metabolism. Uup is a 72?kDa polypeptide which comprises two ABC domains, separated by a 75-residue linker, and a C-terminal domain (CTD) of unknown function. The Uup protein from Escherichia coli has been shown to bind DNA in vitro, and the CTD domain contributes to the DNA-binding affinity. We have produced and purified uniformly labeled ¹⁵N- and ¹⁵N/¹³C Uup CTD domain (region 528?C635), and assigned backbone and side-chains resonances using heteronuclear NMR spectroscopy. Secondary structure evaluation based on backbone chemical shifts is consistent with the presence of three ??-helices, including two long ones (residues 564?C590 and 601?C632), suggesting that Uup CTD may fold as an intramolecular coiled coil motif. This work provides the starting point towards determining the first atomic structure of a non-ATPase domain within the vast REG subfamily of ABC soluble ATPases.     

Microbial Endoliths on East Adriatic Limestone Coast: Morphological vs. Molecular Diversity

For the first time, a combined genotype-phenotype diversity analysis of endolithic microorganisms by pyrosequencing and microscopy was carried out across the intertidal and supratidal ranges of the Adriatic limestone coast. The coastal profiles represent ecotones between the sea and land, which express gradients of water supply, solar illumination, and salinity. The use of scanning electron microscopy imagery of resin casts showed that euendolith penetration remained shallow in the uppermost zones and was progressively deeper in the lower ranges where the water supply was more frequent. Microbial genotypes and phenotypes along the coastal profile showed large differences in dominance of taxa, but less in their diversity. The linkage between morphotypes observed along the profile and sequences was achieved by sequencing single cells/filaments, which were morphologically identified prior to their amplification. Molecular signatures of Hormathonema spp., Hyella caespitosa, Scytonema endolithicum, Solentia paulocellulare and Kyrtuthrix dalmatica were found on the profile and their specific placement was confirmed by morphological observations. Most sequences obtained were affiliated with Cyanobacteria (43%), Alphaproteobacteria (18%), and Gammaproteobacteria (7%). A total of 13 cyanobacterial morphospecies and 17 genotypes were found. Most cyanobacterial sequences belonged to the Pleurocapsales (33%), only 2% of the sequences belonged to the Nostocales and Oscillatoriales. The sequences corresponding to the cyanobacterial genus Hormathonema exhibited the highest abundances among all detected sequences, which is consistent with the microscopic observations in other parts of the world. The results of our investigation underline the importance of a combined approach in comparing the molecular data with microscopic observations in all phases of the study. Further RNA studies are needed to identify the actively growing parts of the endolithic community.     

Coordinating Anions “to the Rescue” of the Lithium Ion Mobility in Ternary Solid Polymer Electrolytes Plasticized With Ionic Liquids

Lithium salts with low coordinating anions such as bis(trifluoromethanesulfonyl)imide (TFSI) have been the state-of-the-art for polyethylene oxide (PEO)-based “dry” polymer electrolytes for 3 decades. Plasticizing PEO with TFSI-based ionic liquids (ILs) to form ternary solid polymer electrolytes (TSPEs) increases conductivity and Li⁺ diffusivity. However, the Li⁺ transport mechanism is unaffected compared to their “dry” counterparts and is essentially coupled to the dynamics of the polymer host matrix, which limits Li⁺ transport improvement. Thus, a paradigm shift is hereby suggested: the utilization of more coordinating anions such as trifluoromethanesulfonyl-N-cyanoamide (TFSAM), able to compete with PEO for Li⁺ solvation, to accelerate the Li⁺ transport and reach a higher Li⁺ transference number. The Li–TFSAM interaction in binary and ternary TFSAM-based electrolytes is probed by experimental methods and discussed in the context of recent computational results. In PEO-based TSPEs, TFSAM drastically accelerates the Li⁺ transport (increases Li⁺ transference number by a factor 6 and the Li⁺ conductivity by 2–3) and computer simulations reveal that lithium dynamics are effectively re-coupled from polymer to anion dynamics. Last, this concept of coordinating anions in TSPEs is successfully applied in LFP||Li metal cells leading to enhanced capacity retention (86% after 300 cycles) and an improved rate performance at 2C.     

How far bowalization affects phytodiversity,life forms and plant morphology in Sub‐humid tropic in West Africa

Bowal or ferricrete, the final of land degradation, occurred only in tropical region. This study aimed at assessing the effects of bowalization on phytodiversity, life forms and morphological response of plant species using Combretum nigricans Leprieur ex Guill. & Perr. as a case study. Morphological parameters (height, number of stems, number of branches, diameter at breast height and crown diameter) of C. nigricans were determined in the sub‐humid zone of Benin. Plant communities were determined according to Multi‐Response Permutation Procedures analysis. Plant communities were more diversified on sand‐clay and concretion soils (control) compared with those described on bowal. C. nigricans developed more stems (3.6 ± 1.4 stems vs. 1.3 ± 0.4 stems), more branches (5.9 ± 2.4 branches vs. 3.2 ± 0.6 branches) and large crown diameter (5 ± 1.48 m vs. 3.4 ± 1.2 m) on bowal than on sand‐clay soil. The best adapted life forms on bowal were therophytes. Bowalization induced loss of phytodiversity, changes in species life forms and provoked local adaptation of tree species.     

Trans-ethnic variation in germline variants of patients with renal cell carcinoma

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Geographical isolation and genetic differentiation: the case of Orestias ascotanensis (Teleostei: Cyprinodontidae), an Andean killifish inhabiting a highland salt pan

Orestias ascotanensis is a killifish endemic to the Ascotán salt pan in the Chilean Altiplano, where it inhabits 12 springs with different degrees of isolation. This species is a suitable model for studying the effect of serial geographical isolations on the differentiation process among populations. The present study examines the genetic variation and structure of the species using mitochondrial DNA control region sequences and eight microsatellite loci, analyzing populations across its distribution range. The evaluation of genetic variation revealed high levels of diversity within the species. The genetic structure analysis showed the existence of four differentiated groups: two groups were formed by the springs located in the northern and southern extremes of the salt pan and two groups were found in the centre of the salt pan. The latter two groups were formed by several springs, most likely as a consequence of the South American summer monsoon that could connect them and allow gene flow. The patterns of genetic differentiation appear to be determined based on the physical isolation of the populations. This isolation may be the result of a combination of factors, including geographical distance, a historical decrease in water levels and altitude differences in the springs of the salt pan. Therefore, this system is a rare example in which hydrological factors can explain genetic differentiation on a very small scale.     

Reply to Hatch and Prihoda

The authors thank Jeff Prince, Ph.D. for his editorial assistance.     

Detection and Capturing of <Superscript>14</Superscript>C Radioactively-Labeled Small Subunit rRNA from Mixed Microbial Communities of a Microbial Mat Using Magnetic Beads

Carbon cycling in the hypersaline microbial mats from Chiprana Lake, Spain is primarily dependent on phototrophic microorganisms with the ability to fix CO₂ into organics that can be further utilized by aerobic as well as anaerobic heterotrophic bacteria. Here, mat pieces were incubated in seawater amended with ¹⁴C sodium bicarbonate and the incorporation of the radiocarbon in the small subunit ribosomal RNA (SSU rRNA) of mat organisms was followed using scintillation counter and autoradiography. Different domains of SSU rRNA were separated from the total RNA by means of streptavidin-coated magnetic beads and biotin-labeled oligonucleotide probes. The ¹⁴C label was detected in isolated RNA by both scintillation counter and autoradiography, however the latter technique was less sensitive. Using scintillation counter, the radiolabel incorporation increased with time with a maximum rate of 0.18 Bq ng⁻¹ detected after 25 days. The bacterial SSU rRNA could be captured using the magnetic beads, however the hybridization efficiency was around 20%. The captured RNA was radioactively labeled, which could be mainly due to the fixation of radiocarbon by phototrophic organisms. In conclusion, the incubation of microbial mats in the presence of radiolabeled bicarbonate leads to the incorporation of the ¹⁴C label into RNA molecules through photosynthesis and this label can be detected using scintillation counter. The used approach could be useful in studying the fate of fixed carbon and its uptake by other microorganisms in complex microbial mats, particularly when species-specific probes are used and the hybridization efficiency and RNA yield are further optimized.     

Sequence Analysis of the Internal Transcribed Spacer 2 (ITS2) from Yeast Species Within the Genus Candida

Nucleotide sequences of the internal transcribed spacer 2 (ITS2) regions were determined for 13 species within the genus Candida, representing a collection of those species pathogenic for humans. No two species had identical sequences and the sizes of ITS2 varied fourfold, representing an apparent continuous gradient of nucleotides. When present, sequence homologies were observed in the 5′ end of ITS2, and many species exhibited more limited homologies within three known conserved domains found in other yeasts. Cluster analysis of primary sequence revealed a concordance with a known taxonomic subfamily and suggests that certain species within the genus form a similar grouping. A majority of species exhibited similar presumptive RNA secondary structures, consistent with the hypothesis that these spacer regions are essential for correct processing of the 5.8S and 28S subunits. Received: 14 April 1997 / Accepted: 22 July 1997