The H-loop in the second nucleotide-binding domain of the cystic fibrosis transmembrane conductance regulator is required for efficient chloride channel closing.

The cystic fibrosis transmembrane conductance regulator (CFTR) is an ATP-binding cassette (ABC) transporter that functions as a cAMP-activated chloride channel. The recent model of CFTR gating predicts that the ATP binding to both nucleotide-binding domains (NBD1 and NBD2) of CFTR is required for the opening of the channel, while the ATP hydrolysis at NBD2 induces subsequent channel closing. In most ABC proteins, efficient hydrolysis of ATP requires the presence of the invariant histidine residue within the H-loop located in the C-terminal part of the NBD. However, the contribution of the corresponding region (H-loop) of NBD2 to the CFTR channel gating has not been examined so far. Here we report that the alanine substitution of the conserved dipeptide HR motif (HR-->AA) in the H-loop of NBD2 leads to prolonged open states of CFTR channel, indicating that the H-loop is required for efficient channel closing. On the other hand, the HR-->AA substitution lead to the substantial decrease of CFTR-mediated current density (pA/pF) in transfected HEK 293 cells, as recorded in the whole-cell patch-clamp analysis. These results suggest that the H-loop of NBD2, apart from being required for CFTR channel closing, may be involved in regulating CFTR trafficking to the cell surface.

Bioelectricity production from wood hydrothermal-treatment wastewater: Enhanced power generation in MFC-fed mixed wastewaters.

Electrogenic microorganisms are the heart of microbial fuel cell (MFC) systems that enable the conversion of waste into bioelectricity. Bacteria able to generate current, found in various natural and anthropogenic environments, need simple substrates such as acetate or glucose. Complex substrates are utilized by bacterial consortia made up of strains that exhibit a wide range of enzymatic and metabolic activity that determines the type of substrate they are able to degrade. The characteristics of the environment that a bacterial consortium develops in strongly affect the consortium's species composition and electrogenic potential. This study presents the first attempt to use industrial raw wastewater from the hydrothermal treatment of wood (WHTW) as a substrate and a source of bacterial consortia for MFC, so that such wastewater could simultaneously be treated and produce bioelectricity. The power generated in MFCs fed with WHTW was enhanced remarkably from 70 to 360mW/m2 when municipal wastewater was introduced into the reactor. An analysis of the bacterial composition of these two types of wastewater revealed that the WHTW was dominated by the genera Thermoanaerobacterium and Paenibacillus while in the biofilm developed in the anode the main genera were Hydrogenophilus and Anaerobaculum. It has been shown for the first time that highly polluted wood industry wastewater may be effectively treated in MFC systems and the use of appropriate bacterial consortium may result in enhancing power generation accompanying wastewater treatment.

Potassium currents in human myogenic cells from donors of different ages.

Ageing in humans is accompanied by a reduction in the capacity of satellite cells to proliferate and the forming myoblasts to fuse. The processes of myoblast differentiation and fusion are associated with specific changes in the cells electrical properties. We wanted to elucidate the possible effects of ageing on these parameters and performed whole-cell patch-clamp recordings on human myoblasts obtained from biopsies of skeletal muscles from 2-, 48- and 76-year-old donors. First, we found that resting membrane potential on the 4th day of differentiation in vitro is less negative in the older than in the younger cells. Moreover, the oldest cells showed a smaller density of outward and inward potassium currents. More cells from the old and middle-age donors have a low (less than -40 mV) potential of activation for the outward potassium current. We conclude that in human myoblasts biophysical properties of potassium currents change with donor age.

Design and characterization of novel all-solid-state potentiometric sensor array dedicated to physiological measurements.

A novel construction of all-solid-state potentiometric sensor array designed for physiological measurements has been presented. The planar construction and elimination of liquid phase creates broad opportunities for the modifications in the sensing part of the sensor. The designed construction is based on all-solid-state ion-selective electrodes integrated with the ionic-liquid based reference electrode. Work parameters of the sensor arrays were characterized. It has been shown that presented sensor design indicates high sensitivity (55.2±1mV/dec, 56.3±2mV/dec, 58.4±1mV/dec and 53.5±1mV/pH for sodium-, potassium-, chloride- and pH-selective electrodes, respectively in 10(-5)-10(-1.5)M range of primary ions), low response time (t95 did not exceed 10s), high potential stability (potential drift in 28-h measurement was ca. ±2mV) and potential repeatability ca. ±1mV. The system was successfully applied to the simultaneous determination of K(+), Cl(-), Na(+) and pH in the model physiological solution and for the ion flux studies in human colon epithelium Caco-2 cell line as well.