The fourth extracellular loop of the alpha subunit of Na,K-ATPase. Functional evidence for close proximity with the second extracellular loop.

Na,K-ATPase is the main active transport system that maintains the large gradients of Na(+) and K(+) across the plasma membrane of animal cells. The crystal structure of a K(+)-occluding conformation of this protein has been recently published, but the movements of its different domains allowing for the cation pumping mechanism are not yet known. The structure of many more conformations is known for the related calcium ATPase SERCA, but the reliability of homology modeling is poor for several domains with low sequence identity, in particular the extracellular loops. To better define the structure of the large fourth extracellular loop between the seventh and eighth transmembrane segments of the alpha subunit, we have studied the formation of a disulfide bond between pairs of cysteine residues introduced by site-directed mutagenesis in the second and the fourth extracellular loop. We found a specific pair of cysteine positions (Y308C and D884C) for which extracellular treatment with an oxidizing agent inhibited the Na,K pump function, which could be rapidly restored by a reducing agent. The formation of the disulfide bond occurred preferentially under the E2-P conformation of Na,K-ATPase, in the absence of extracellular cations. Using recently published crystal structure and a distance constraint reproducing the existence of disulfide bond, we performed an extensive conformational space search using simulated annealing and showed that the Tyr(308) and Asp(884) residues can be in close proximity, and simultaneously, the SYGQ motif of the fourth extracellular loop, known to interact with the extracellular domain of the beta subunit, can be exposed to the exterior of the protein and can easily interact with the beta subunit.

Why did Swiss citizens refuse a comprehensive second-hand smoke ban?

The ill effects of second-hand smoke are now well documented. To protect the population from exposure to tobacco smoke, comprehensive smoking bans are necessary as expressed in the WHO Framework Convention on Tobacco Control and its guidelines. Switzerland has only a partial smoking ban full of exceptions which has been in effect since 2010, which reproduces the so-called Spanish model. In September 2012, the Swiss citizens refused a proposal for a more comprehensive ban. This case study examines the reasons behind this rejection and draws some lessons that can be learnt from it.

Exploring the relative DNA contribution of first and second object's users on mock touch DNA mixtures

Contact stains recovered at break-in crime scenes are frequently characterized by mixtures of DNA from several persons. Broad knowledge on the relative contribution of DNA left behind by different users overtime is of paramount importance. Such information might help crime investigators to robustly evaluate the possibility of detecting a specific (or known) individual's DNA profile based on the type and history of an object. To address this issue, a contact stain simulation-based protocol was designed. Fourteen volunteers either acting as first or second object's users were recruited. The first user was required to regularly handle/wear 9 different items during an 8-10-day period, whilst the second user for 5, 30 and 120 min, in three independent simulation sessions producing a total of 231 stains. Subsequently, the relative DNA profile contribution of each individual pair was investigated. Preliminary results showed a progressive increase of the percentage contribution of the second user compared to the first. Interestingly, the second user generally became the major DNA contributor when most objects were handled/worn for 120 min, Furthermore, the observation of unexpected additional alleles will then prompt the investigation of indirect DNA transfer events.