Search

Fas apoptosis inhibitory molecule (FAIM) is a soluble cytosolic protein inhibitor of programmed cell death and is found in organisms throughout the animal kingdom. A short isoform of FAIM is expressed in all tissue types, while an alternatively spliced long isoform is specifically expressed in the brain. Here, the short isoform is shown to consist of two independently folding domains in contact with each other. The NMR solution structure of the C-terminal domain of murine FAIM is solved in isolation and revealed to be a novel protein fold, a noninterleaved seven-stranded beta-sandwich. The structure and sequence reveal several residues that are likely to be involved in functionally significant interactions with the N-terminal domain or other binding partners. Chemical shift perturbation is used to elucidate contacts made between the N-terminal domain and the C-terminal domain.

Two mass spectrometers (MS) are tested for the measurement of volatile substances, such as hydrocarbons and metabolic gases, in natural waters. KOALA is a backpackable MS operated from above the water surface, in which samples are pumped through a flow cell using a syringe. NEREUS is an underwater instrument hosted by an autonomous underwater vehicle (AUV) that is linked to a communications network to provide chemical data in real time. The mass analyzers of the two MS are nearly identical cycloids, and both use flat-plate membrane inlets. Testing took place in an eutrophic, thermally stratified lake exhibiting steep chemical gradients and significant levels of methane. KOALA provided rapid multispecies analysis of dissolved gases, with a detection limit for methane of 0.1 ppm (readily extendable to 0.01 ppm) and savings of time of at least a factor of 10 compared to that of conventional analysis. The AUV-mounted NEREUS additionally provided rapid spatial coverage and the capability of performing chemical surveys autonomously. Tests demonstrated the need for temperature control of a membrane inlet when steep thermal gradients are present in a water body, as well as the benefits of co-locating all sensors on the AUV to avoid interference from chemically different waters entering and draining from the free-flooding outer hull. The ability to measure dissolved volatiles provided by MS offers potential for complementarity with ionic sensors in the study of natural waters, such as in the case of the carbonate system.

The dsRNA-dependent protein kinase (PKR) is a key mediator of the anti-viral and anti-proliferative effects of interferon. Unphosphorylated PKR is characterized by inhibitory interactions between the kinase and RNA binding domains (RBDs), but the structural details of the latent state and its unraveling during activation are not well understood. To study PKR regulation by NMR we assigned a large portion of the backbone resonances of the catalytically inactive K296R kinase domain, and performed (15)N-heteronuclear single quantum coherence (HSQC) titrations of this kinase domain with the RBDs. Chemical shift perturbations in the kinase indicate that RBD2 binds to the substrate eIF2alpha docking site in the kinase C-lobe. Consistent with these results, a mutation in the eIF2alpha docking site, F495A, displays weaker interactions with the RBD. The full-length RBD1+2 binds more strongly to the kinase domain than RBD2 alone. The observed chemical shift changes extend from the eIF2alpha binding site into the kinase N-lobe and inside the active site, consistent with weak interactions between the N-terminal part of the RBD and the kinase.

Vasopressin (VS) has been reported to stimulate colonic peristalsis in different therapeutic conditions. In order to determine the mechanisms involved in this effect, colonic function was studied with three different techniques staying: a) the transit time of radioopaque markers through the colon was measured in 7 healthy subjects after VS IM (0.3 U/kg of weight). A marked propulsive effect was observed. One hour after injection, 64.7 +/- 16.2 p. 100 of the markers (m +/- sem) had left the right colon vs 9.1 +/- 4.6 p. 100 after injection of NaCl (p less than 0.01) and 70.5 +/- 10.8 p. 100 of the markers emptied from the left colon vs 4.3 +/- 4.5 p. 100 after NaCl (p less than 0.01); b) an electromyographic study was carried out in 6 other healthy subjects with an intraluminal device equipped with contact electrodes, introduced into the left colon by colonoscopy. The injection of VS was followed by an increase in the number of the propagating electrical spike bursts that are known to correlate well with the propulsion of the colonic intraluminal contents. The number of bursts was 2.7 +/- 0.6 bursts/30 min after NaCl and 5.2 +/- 1.4 bursts/30 min after VS (p less than 0.02); c) finally, the outflow of ileostomies and colostomies was measured in respectively 3 and 8 subjects over one h after IM 0.3 U/kg of VS. A considerable increase in the outflow of intestinal fluids was observed: output from the colostomies was 10 +/- 10 ml/h after NaCl and 250 +/- 39 ml/h after VS (p less than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)