Comprehensive micropollutant screening using LC-HRMS/MS at three riverbank filtration sites to assess natural attenuation and potential implications for human health.

Riverbank filtration (RBF) is used worldwide to produce high quality drinking water. With river water often contaminated by micropollutants (MPs) from various sources, this study addresses the occurrence and fate of such MPs at three different RBF sites with oxic alluvial sediments and short travel times to the drinking water well down to hours. A broad range of MPs with various physico-chemical properties were analysed with detection limits in the low ng L range using solid phase extraction followed by liquid chromatography coupled to tandem high resolution mass spectrometry.

The thermal impact of subsurface building structures on urban groundwater resources - A paradigmatic example.

Shallow subsurface thermal regimes in urban areas are increasingly impacted by anthropogenic activities, which include infrastructure development like underground traffic lines as well as industrial and residential subsurface buildings. In combination with the progressive use of shallow geothermal energy systems, this results in the so-called subsurface urban heat island effect. This article emphasizes the importance of considering the thermal impact of subsurface structures, which commonly is underestimated due to missing information and of reliable subsurface temperature data.

Photo-redox reactions of dicarboxylates and α-hydroxydicarboxylates at the surface of Fe(III)(hydr)oxides followed with in situ ATR-FTIR spectroscopy.

Colloidal mineral-phases play an important role in the adsorption, transport and transformation of organic and inorganic compounds in the atmosphere and in aqueous environments. Artificial UV-light and sunlight can induce electron transfer reactions between metal ions of the solid phases and adsorbed compounds, leading to their transformation and degradation. To investigate different possible photo-induced oxidation pathways of dicarboxylates adsorbed on iron(III)(hydr)oxide surfaces, we followed UV-A induced photoreactions of oxalate, malonate, succinate and their corresponding α-hydroxy analogues tartronate and malate with in situ ATR-FTIR spectroscopy in immersed particle layers of lepidocrocite, goethite, maghemite and hematite at pH 4.

Probing the RNA binding surface of the HIV-1 nucleocapsid protein by site-directed mutagenesis.

The highly conserved nucleocapsid protein domain in HIV-1 recognizes and binds SL3 in genomic RNA. In this work, we used the structure of the NCp7-SL3 RNA complex to guide the construction of 16 NCp7 mutants to probe the RNA binding surface of the protein [De Guzman, R. N.

Sampling a biomarker of the human immunodeficiency virus across a synthetic nanopore.

One primary goal in nanobiotechnology is designing new methodologies for molecular biomedical diagnosis at stages much earlier than currently possible and without use of expensive reagents and sophisticated equipment. In this work, we show the proof of principle for single-molecule detection of the nucleocapsid protein 7 (NCp7), a protein biomarker of the HIV-1 virus, using synthetic nanopores and the resistive-pulse technique. The biosensing mechanism relied upon specific interactions between NCp7 and aptamers of stem-loop 3 (SL3) in the packaging domain of the retroviral RNA genome.

Engineering a rigid protein tunnel for biomolecular detection.

One intimidating challenge in protein nanopore-based technologies is designing robust protein scaffolds that remain functionally intact under a broad spectrum of detection conditions. Here, we show that an extensively engineered bacterial ferric hydroxamate uptake component A (FhuA), a β-barrel membrane protein, functions as a robust protein tunnel for the sampling of biomolecular events. The key implementation in this work was the coupling of direct genetic engineering with a refolding approach to produce an unusually stable protein nanopore.

The Response of Shewanella oneidensis MR-1 to Cr(III) Toxicity Differs from that to Cr(VI).

Chromium is a contaminant of concern that is found in drinking water in its soluble, hexavalent form [Cr(VI)] and that is known to be toxic to eukaryotes and prokaryotes. Trivalent chromium [Cr(III)] is thought to be largely harmless due to its low solubility and inability to enter cells. Previous work has suggested that Cr(III) may also be toxic to microorganisms but the mechanism remained elusive.

Acyclic identification of aptamers for human alpha-thrombin using over-represented libraries and deep sequencing.

Background: Aptamers are oligonucleotides that bind proteins and other targets with high affinity and selectivity. Twenty years ago elements of natural selection were adapted to in vitro selection in order to distinguish aptamers among randomized sequence libraries. The primary bottleneck in traditional aptamer discovery is multiple cycles of in vitro evolution.

Effects of the nature and concentration of salt on the interaction of the HIV-1 nucleocapsid protein with SL3 RNA.

The mature nucleocapsid protein of HIV-1, NCp7, and the NC domains in gag precursors are attractive targets for anti-AIDS drug discovery. The stability of the 1:1 complex of NCp7 with a 20mer mimic of stem-loop 3 RNA (SL3, also called psi-RNA, in the packaging domain of genomic RNA) is strongly affected by changes in ionic strength. NC domains recognize and specifically package genomic HIV-1 RNA, while electrostatic attractions and high concentrations of protein and RNA drive NCp7 to completely coat the RNA in the mature virion.

Wavelength-dependence of photoreductive dissolution of lepidocrocite (gamma-FeOOH) in the absence and presence of the siderophore DFOB.

Photoreductive dissolution of lepidocrocite (gamma-FeOOH) in the presence/absence of the siderophore desferrioxamine B (DFOB) was investigated at different wavelengths. At pH 3 in the absence of DFOB, Fe(II) formation rates normalized to the photon flux increased with decreasing wavelengths below 515 nm, consistent with enhanced Fe(II) formation at lower wavelengths by photolysis of surface Fe(III)-hydroxo groups or by surface scavenging of photoelectrons generated in the semiconducting bulk. In the presence of DFOB at pH 3, photoreductive dissolution rates, normalized to the photon flux, increased more strongly with decreasing wavelengths below 440 nm.

Photoreductive dissolution of iron(III) (hydr)oxides in the absence and presence of organic ligands: experimental studies and kinetic modeling.

This study investigated the kinetics of the photoreductive dissolution of various iron(III)(hydr)oxide phases, lepidocrocite (gamma-FeOOH), ferrihydrite, and hydrous ferric oxide, in the absence of organic ligands as a function of pH in deaerated and aerated suspensions. Photoreductive dissolution of lepidocrocite and ferrihydrite only occurred below pH 6. Under oxic conditions, we observed both the formation of aqueous Fe(II) and H2O2 during photoreductive dissolution of lepidocrocite and ferrihydrite at pH 3.

A nucleic acid switch triggered by the HIV-1 nucleocapsid protein.

A unimolecular oligonucleotide switch, termed here an AlloSwitch, binds the mature HIV-1 nucleocapsid protein, NCp7. This switch can be used as an indicator for the presence of free NCp7 and NC domains in precursor and fusion proteins. It is thermodynamically stable in two conformations, H and O.

Iron isotope fractionation during proton-promoted, ligand-controlled, and reductive dissolution of Goethite.

Iron isotope fractionation during dissolution of goethite (alpha-FeOOH) was studied in laboratory batch experiments. Proton-promoted (HCl), ligand-controlled (oxalate dark), and reductive (oxalate light) dissolution mechanisms were compared in order to understand the behavior of iron isotopes during natural weathering reactions. Multicollector ICP-MS was used to measure iron isotope ratios of dissolved iron in solution.

Changes in drug 13C NMR chemical shifts as a tool for monitoring interactions with DNA.

The antibiotic drug, netropsin, was complexed with the DNA oligonucleotide duplex [d(GGTATACC)]2 to monitor drug 13C NMR chemical shifts changes. The binding mode of netropsin to the minor groove of DNA is well-known, and served as a good model for evaluating the relative sensitivity of 13C chemical shifts to hydrogen bonding. Large downfield shifts were observed for four resonances of carbons that neighbor sites which are known to form hydrogen bond interactions with the DNA minor groove.

Revised UV extinction coefficients for nucleoside-5'-monophosphates and unpaired DNA and RNA.

Ultraviolet absorption provides the nearly universal basis for determining concentrations of nucleic acids. Values for the UV extinction coefficients of DNA and RNA rely on the mononucleotide values determined 30-50 years ago. We show that nearly all of the previously published extinction coefficients for the nucleoside-5'-monophosphates are too large, and in error by as much as 7%.

Stem of SL1 RNA in HIV-1: structure and nucleocapsid protein binding for a 1 x 3 internal loop.

The 5'-leader of HIV-1 RNA controls many viral functions. Nucleocapsid (NC) domains of gag-precursor proteins select genomic RNA for packaging by binding several sites in the leader. One is likely to be a stem defect in SL1 that can adopt either a 1 x 3 internal loop, SL1i (including G247, A271, G272, G273) or a 1 x 1 internal loop (G247 x G273) near a two-base bulge (A269-G270).

Affinities of the nucleocapsid protein for variants of SL3 RNA in HIV-1.

Efficient packaging of genomic RNA into new HIV-1 virus particles requires that nucleocapsid domains of precursor proteins bind the SL3 tetraloop (G317-G-A-G320) from the 5'-untranslated region. This paper presents the affinities of 35 RNA variants of SL3 for the mature 55mer NC protein, as measured by fluorescence quenching of tryptophan-37 in the protein by nucleobases. The 1:1 complexes that form in 0.

Accurate nucleic acid concentrations by nuclear magnetic resonance.

Determination of the concentration of biochemical samples often yields values with uncertainties of 10-20% or more. This paper details a protocol for use with 500- to 600-MHz NMR spectrometers to measure approximately 1mM concentrations within +/-1-3% accuracy. With suitable precautions, all compounds have equal NMR "absorption coefficients" for protons.

Affinities of packaging domain loops in HIV-1 RNA for the nucleocapsid protein.

To design anti-nucleocapsid drugs, it is useful to know the affinities the protein has for its natural substrates under physiological conditions. Dissociation equilibrium constants are reported for seven RNA stem-loops bound to the mature HIV-1 nucleocapsid protein, NCp7. The loops include SL1, SL2, SL3, and SL4 from the major packaging domain of genomic RNA.

Regioselective synthesis of 1,3,5-13C3 and 2,4-13C2-labeled 2-deoxyribonolactones.

The synthesis of 1,3,5-13C3- and 2,4-13C2-labeled 5-O-bromobenzyl-2-deoxyribonolactones 2, precursors to 13C-enriched nucleoside phosphoramidites for solid-phase synthesis of DNA oligonucletides, is described. An equimolar combination of these two multiply labeled lactones affords a "population-labeled" mixture of isotopomers which exhibits an approximately 50-fold increase in the sensitivity of 13C-NMR compared to natural abundance measurements. The 13C-13C 2-bond and 4-bond coupling constants are reported for the lactones; all are <2Hz, confirming that this labeling scheme should be especially useful for NMR-relaxation measurements.

Structure of SL4 RNA from the HIV-1 packaging signal.

The NMR-based structure is described for an RNA model of stem-loop 4 (SL4) from the HIV-1 major packaging domain. The GAGA tetraloop adopts a conformation similar to the classic GNRA form, although there are differences in the details. The type II tandem G.

Changes in 13C NMR chemical shifts of DNA as a tool for monitoring drug interactions.

The antibiotic drug, netropsin, was complexed with the DNA oligonucleotide duplex [d(GGTATACC)]2 to explore the effects of ligand binding on the 13C NMR chemical shifts of the DNA base and sugar carbons. The binding mode of netrospin to TA-rich tracts of DNA has been well documented and served as an attractive model system. For the base carbons, four large changes in resonance chemical shifts were observed upon complex formation: -0.

Three-dimensional folding of an RNA hairpin required for packaging HIV-1.

An NMR-based structure is presented for a 20 mer hairpin model of the SL3 stem-loop from the HIV-1 packaging signal. The stem has an A-family structure. However, the GGAG tetraloop appears to be flexible with the second (G10) and fourth (G12) bases extruded from the normal stacking arrangement.

Full 1H NMR assignment of a 24-nucleotide RNA hairpin: application of the 1H 3D-NOE/2QC experiment.

The subject RNA models the binding site for the coat protein of the R17 virus, as well as the ribosome recognition sequence for the R17 replicase gene. With an RNA of this size, overlaps among the sugar protons complicate assignments of the 1H NMR spectrum. The cross peaks that overlap significantly in 2D-NOE spectra can frequently be resolved by introducing a third, in our approach the double-quantum, frequency axis.