Intramolecular photoinduced electron transfer in zwitterionic quinolinium dyes - Experimental and theoretical studies

Quinolinium cations and quinolinium betaines were investigated in the representative solvents water and acetonitrile at room temperature using stationary and time-resolved fluorescence spectroscopy (Single-Photon-Counting-method). Experimental results reveal that sulfoalkyl- and carboxyalkyl-quinolinium compounds display a strikingly different behavior in the two solvents. Furthermore, the fluorescence lifetime depends on the length of the spacer for the sulfoalkyl compounds in acetonitrile and the carboxyalkyl compounds in water, respectively. This suggests an intramolecular interaction of the anionic headgroups with the quinolinium system in the excited state. To support this idea. different positions at the chromophore are substituted by a methylgroup in order to perturb the proposed interaction. With the intention to understand the dynamics of the postulated photoinduced electron transfer from the anionic group onto the excited quinolinium chromophore, semiempirical quantum chemical calculations were performed on the species using the PM3 hamiltonian including solvent effects by a self consistent reaction field (SCRF). We show that the Marcus theory of electron transfer may serve as a theoretical basis for a natural interpretation of the dynamic fluorescence quenching behavior

Microscopy of bacterial translocation during small bowel obstruction and ischemia in vivo – a new animal model

BACKGROUND: Existing animal models provide only indirect information about the pathogenesis of infections caused by indigenous gastrointestinal microflora and the kinetics of bacterial translocation. The aim of this study was to develop a novel animal model to assess bacterial translocation and intestinal barrier function in vivo. METHODS: In anaesthetized male Wistar rats, 0.5 ml of a suspension of green fluorescent protein-transfected E. coli was administered by intraluminal injection in a model of small bowel obstruction. Animals were randomly subjected to non-ischemic or ischemic bowel obstruction. Ischemia was induced by selective clamping of the terminal mesenteric vessels feeding the obstructed bowel loop. Time intervals necessary for translocation of E. coli into the submucosal stroma and the muscularis propria was assessed using intravital microscopy. RESULTS: Bacterial translocation into the submucosa and muscularis propria took a mean of 36 ± 8 min and 80 ± 10 min, respectively, in small bowel obstruction. Intestinal ischemia significantly accelerated bacterial translocation into the submucosa (11 ± 5 min, p < 0.0001) and muscularis (66 ± 7 min; p = 0.004). Green fluorescent protein-transfected E. coli were visible in frozen sections of small bowel, mesentery, liver and spleen taken two hours after E. coli administration. CONCLUSIONS: Intravital microscopy of fluorescent bacteria is a novel approach to study bacterial translocation in vivo. We have applied this technique to define minimal bacterial transit time as a functional parameter of intestinal barrier function

Active-state models of ternary GPCR complexes: determinants of selective receptor-G-protein coupling

Based on the recently described crystal structure of the β2 adrenergic receptor--Gs-protein complex, we report the first molecular-dynamics simulations of ternary GPCR complexes designed to identify the selectivity determinants for receptor-G-protein binding. Long-term molecular dynamics simulations of agonist-bound β2AR-Gαs and D2R-Gαi complexes embedded in a hydrated bilayer environment and computational alanine-scanning mutagenesis identified distinct residues of the N-terminal region of intracellular loop 3 to be crucial for coupling selectivity. Within the G-protein, specific amino acids of the α5-helix, the C-terminus of the Gα-subunit and the regions around αN-β1 and α4-β6 were found to determine receptor recognition. Knowledge of these determinants of receptor-G-protein binding selectivity is essential for designing drugs that target specific receptor/G-protein combinations

Molecular dynamics simulation links conformation of a pore-flanking region to hyperekplexia-related dysfunction of the inhibitory glycine receptor.

AbstractInhibitory glycine receptors mediate rapid synaptic inhibition in mammalian spinal cord and brainstem. The previously identified hyperekplexia mutation GLRA1(P250T), located within the intracellular TM1-2 loop of the GlyR α1 subunit, results in altered receptor activation and desensitization. Here, elementary steps of ion channel function of α1(250) mutants were resolved and shown to correlate with hydropathy and molar volume of residue α1(250). Single-channel recordings and rapid activation kinetic studies using laser pulse photolysis showed reduced conductance but similar open probability of α1(P250T) mutant channels. Molecular dynamics simulation of a helix-turn-helix motif representing the intracellular TM1-2 domain revealed alterations in backbone conformation, indicating an increased flexibility in these mutants that paralleled changes in elementary steps of channel function. Thus, the architecture of the TM1-2 loop is a critical determinant of ion channel conductance and receptor desensitization