Production, extraction, purification, and stabilization of integral membrane proteins are key steps for successful structural biology studies, in particular for X-ray crystallography or single particle microscopy. Here, we present the purification protocol of CntI from Pseudomonas aeruginosa, a new metallophore exporter of the Drug Metabolite Transporter (DMT) family involved in pseudopaline secretion. Subsequent to CntI purification, we optimized the buffer pH, salts, and additives by differential scanning fluorimetry (DSF), also known as Thermofluor Assay (TFA) or fluorescent thermal stability assay (FTSA), with the use of dye 1-AnilinoNaphthalene-8-Sulfonic acid (ANS), a fluorescent molecule compatible with detergents. After the buffer optimization, the purified CntI was analyzed by Size Exclusion Chromatography coupled with Multi-Angle Laser Light Scattering (SEC-MALLS), UV absorbance, and Refractive Index detectors, in order to determine the absolute molar mass of the protein-detergent complex, the detergent amount bound to the protein and the amount of protein-free detergent micelles. Altogether, these biophysical techniques give preliminary and mandatory information about the suitability of the purified membrane protein for further biophysical or structural investigations.

Leafy vegetables are used in various cuisines worldwide; however, as they cannot be peeled and their leaf surface area is large, the risk of retaining pesticide residues on these vegetables is relatively higher than on others. To our knowledge, this is the first comparative study to reveal the effect of removing pesticide residues from five artificially contaminated leafy vegetables (lettuce, perilla leaves, spinach, crown daisy, and ssamchoo (Brassica lee ssp. namai)) using different removal methods. The percent reduction range for each method was 43.7−77.0%, and the reduction range for the five leafy vegetables was 40.6−67.4%. Lettuce had the highest reduction (67.4 ± 7.3%), whereas ssamchoo had the lowest reduction (40.6 ± 12.9%). Spinach and crown daisy showed no significant difference in their reductions. Based on reduction by method, running water (77.0 ± 18.0%) and boiling (59.5 ± 31.2%) led to the highest reduction, whereas detergent (43.7 ± 14.5%) led to the lowest reduction. The reductions of chlorfenapyr, diniconazole, indoxacarb, fludioxonil, pyraclostrobin, and lufenuron in the leafy vegetables were lower with blanching and boiling than with other methods (p < 0.05). These results highlight the importance of thoroughly washing leafy vegetables to lower the intake of pesticide residues before cooking.

Surfactants play a very important role in laundry and household cleaning products ingredients. In this research, the application of lipopeptide biosurfactants, produced by Bacillus subtilis SPB1, in the formulation of a washing powder was investigated. The SPB1 biosurfactant was mixed with sodium tripolyphosphate as a builder and sodium sulfate as filler. The efficiency of the formulated detergent composition with different washing conditions to remove a stain from cotton fabric was examined. The results showed that the formulated detergent was effective in oil removal, with optimal washing conditions of pH, temperature, striate and time of washing system of 7, 65°C, 1000 RPM and 60 min, respectively. A comparative study of different detergent compositions (biosurfactant-based detergent, combined biosurfactant-commercial detergent, and a commercial detergent) for the removal of oil and tea stains, proved that the bio-scouring was more effective (>75%) in terms of the stain removal than the commercial powders (<60%). Moreover, the results demonstrated that the biosurfactant acts additively with a commercial detergent and enhances their performance from 33 to 45% in removing oil stain and from 57 to 64% in removing tea stain. As a conclusion, in addition to the low toxicity and the high biodegradability of the microbial biosurfactants, the results of this study have shown that the future use of this lipopeptide biosurfactant as laundry detergent additive is highly promising.

Solid-state NMR spectroscopy has been developed for the investigation of membrane-associated polypeptides and remains one of the few techniques to reveal high-resolution structural information in liquid-disordered phospholipid bilayers. In particular, oriented samples have been used to investigate the structure, dynamics and topology of membrane polypeptides. Much of the previous solid-state NMR work has been developed and performed on peptides but the technique is constantly expanding towards larger membrane proteins. Here, a number of protocols are presented describing among other the reconstitution of membrane proteins into oriented membranes, monitoring membrane alignment by 31P solid-state NMR spectroscopy, investigations of the protein by one- and two-dimensional 15N solid-state NMR and measurements of the lipid order parameters using 2H solid-state NMR spectroscopy. Using such methods solid-state NMR spectroscopy has revealed a detailed picture of the ensemble of both lipids and proteins and their mutual interdependence in the bilayer environment.

BACKGROUND: The sensitivity of the commonly used immunochromatographic fecal occult blood test (iFOBT) is critical in the routine screening of digestive tract cancers.
OBJECTIVE: To determine whether the performance of the iFOBT might be improved by optimizing detergents and buffers in stool solution.
METHODS: We tested different buffers and detergents in stool solution. The specificity of the new solution condition was confirmed by dot immunoblotting. A total of 1122 clinical stool specimens were collected to evaluate the clinical application of this refined stool solution.
RESULTS: The different salts exerted minor effects; also, sodium dodecyl sulfate (SDS) prevented the test from working as designed. However, 1% Triton X-100 improved the sensitivity significantly, as confirmed by dot immunoblot testing. With 1% Triton X-100 in the stool solution, the rate of positive iFOBT results in patients with digestive tract cancers increased, from 40.4% to 49.4%.
CONCLUSIONS: Use of 1% Triton X-100 improves the sensitivity of the iFOBT and thus, this substance should be routinely included in iFOBT for screening of digestive tract cancers.

Small angle neutron scattering (SANS) is a powerful tool to obtain structural information on solubilized membrane proteins on the nanometer length-scale in complement to other structural biology techniques such as cryo-EM, NMR and SAXS. In combination with deuteration of components and/or contrast variation (H2O:D2O exchange in the buffer) SANS allows to separate structural information from the protein and the detergent/lipid parts in solution. After a short historical overview on results obtained by SANS on membrane protein systems, this book chapter introduces the basic theoretical principles of the technique as well as requirements on samples. The two introductory sections are followed by an illustration of the specific consequences of sample heterogeneity of solubilized membrane proteins in the presence of detergent/lipid molecules on the interpretation of structural information by using simple, geometric models. The next sections deal with more sophisticated modelling approaches including ab initio shape reconstructions and full-atomic models in the presence of detergent/lipid and specific results obtained by these approaches. After a short comparison with the SAXS technique, this book chapter concludes with an overview of present and future developments and impact that can be expected by SANS on membrane structural biology in the coming years.

Privacy curtains, frequently used in hospitals to separate patient care areas may have an important role in the transmission of healthcare-associated pathogens. In this pilot study, we inoculated curtain swatches with suspensions of clinical specimens of meticillin resistant Staphylococcus aureus (MRSA), vancomycin-resistant enterococcus (VRE), and Clostridium difficile before using a gloved hand to touch the inoculated curtain swatch and transfer to clean agar plates. Three different commonly used disinfectants were then sprayed onto these swatches before using a clean gloved hand to touch the swatch and transfer onto new agar plates. All plates were incubated at 35°C for 24 and 72 h. Bacterial growth before and after disinfection was assessed and compared. 3.1% hydrogen peroxide effectively eliminated transfer of C. difficile, MRSA and VRE from inoculated curtains.

BACKGROUND: The smear layer has the capability to protect the bacteria within the dentinal tubules from intracanal medicament. After removal of the smear layer from infected root canals, it allows disinfection of the entire root canal. The smear layer compromising the seal between the root canal sealer and root canal wall also decreases the penetration of irrigants into dentinal tubules.
OBJECTIVE: This study compares the amount of phosphorous liberated and demineralization of the radicular dentin with 17% ethylenediaminetetraacetic acid, 10% citric acid and mixture of doxycycline, citric acid, and a detergent at different time intervals.
METHODS: Extracted maxillary single-rooted teeth were prepared by using a combination of passive step-back and rotary 0.04 taper nickel-titanium files. Sodium hypochlorite 5.25% and sterile distilled water were used as an intracanal irrigant. The canals were then treated with 5 mL of one of the following solutions such as final rinse sterile distilled water, 5.25% sodium hypochlorite, 17% ethylenediaminetetraacetic acid (EDTA) or mixture of doxycycline, citric acid, and a detergent. The presence or absence of smear layer and the amount of erosion on the surface of the root canal walls at the coronal, middle, and apical portions of each canal were examined under a scanning electron microscope.
METHODS: Data were analyzed by one-way analysis of variance (ANOVA) to determine whether there were significant differences between the groups.
RESULTS: The results show that mixture of doxycycline, citric acid, and a detergent is an effective solution for the removal of the smear layer and does not significantly change the structure of the dentinal tubules.
CONCLUSIONS: In this study, 10% citric acid shows the maximum amount of dimeneralization of radicular dentine followed by mixture of doxycycline, citric acid, and a detergent, and 17% ethylenediaminetetraacetic acid. When all the subgroups were compared, it was seen that the maximum amount of phosphorus liberation was performed by 10% citric acid >mixture of doxycycline, citric acid, and a detergent >17% EDTA at a different time interval.

Ion-coupled secondary transport is utilized by multiple integral membrane proteins as a means of achieving the thermodynamically unfavorable translocation of solute molecules across the lipid bilayer. The chemical nature of these molecules is diverse and includes sugars, amino acids, neurotransmitters, and other ions. LeuT is a sodium-coupled, nonpolar amino acid symporter and eubacterial member of the solute carrier 6 (SLC6) family of Na(+)/Cl(-)-dependent neurotransmitter transporters. Eukaryotic counterparts encompass the clinically and pharmacologically significant transporters for γ-aminobutyric acid (GABA), glycine, serotonin (5-hydroxytryptamine, 5-HT), dopamine (DA), and norepinephrine (NE). Since the crystal structure of LeuT was first solved in 2005, subsequent crystallographic, binding, flux, and spectroscopic studies, complemented with homology modeling and molecular dynamic simulations, have allowed this protein to emerge as a remarkable mechanistic paradigm for both the SLC6 class as well as several other sequence-unrelated SLCs whose members possess astonishingly similar architectures. Despite yielding groundbreaking conceptual advances, this vast treasure trove of data has also been the source of contentious hypotheses. This chapter will present a historical scientific overview of SLC6s; recount how the initial and subsequent LeuT structures were solved, describing the insights they each provided; detail the accompanying functional techniques, emphasizing how they either supported or refuted the static crystallographic data; and assemble these individual findings into a mechanism of transport and inhibition.

Uptake and depuration kinetics of dissolved [(14)C]C₁₂-6-linear alkylbenzene sulfonate (LAS) were determined in the shrimp Palaemonetes varians using environmentally relevant exposure concentration. The shrimp concentrated LAS from seawater with a mean BCF value of 120 L kg(-1) after a 7-day exposure. Uptake biokinetics were best described by a saturation model, with an estimated BCFss, of 159 ± 34 L kg(-1), reached after 11.5 days. Shrimp weight influenced significantly BCF value with smaller individuals presenting higher affinity to LAS. To the light of a whole body autoradiography, major accumulation of LAS occurred in the cephalothorax circulatory system (gills, heart, hepatopancreas) and ocular peduncle, but not in the flesh, limiting potential transfer to human consumers. LAS depuration rate constant value of the shrimp was 1.18 ± 0.08 d(-1) leading to less than 1% of remaining LAS in its tissues after 8 days of depuration.