A G protein gamma subunit-like domain shared between RGS11 and other RGS proteins specifies binding to Gbeta5 subunits

Regulators of G protein signaling (RGS) proteins act as GTPase-activating proteins (GAPs) toward the alpha subunits of heterotrimeric, signal-transducing G proteins. RGS11 contains a G protein gamma subunit-like (GGL) domain between its Dishevelled/Egl-10/Pleckstrin and RGS domains. GGL domains are also found in RGS6, RGS7, RGS9, and the Caenorhabditis elegans protein EGL-10. Coexpression of RGS11 with different Gbeta subunits reveals specific interaction between RGS11 and Gbeta5. The expression of mRNA for RGS11 and Gbeta5 in human tissues overlaps. The Gbeta5/RGS11 heterodimer acts as a GAP on Galphao, apparently selectively. RGS proteins that contain GGL domains appear to act as GAPs for Galpha proteins and form complexes with specific Gbeta subunits, adding to the combinatorial complexity of G protein-mediated signaling pathways.     

Structure and stability of an immunoglobulin superfamily domain from twitchin, a muscle protein of the nematode Caenorhabditis elegans

The NMR solution structure of an immunoglobulin superfamily module of twitchin (Ig 18') has been determined and the kinetic and equilibrium folding behaviour characterised. Thirty molecular coordinates were calculated using a hybrid distance geometry-simulated annealing protocol based on 1207 distance and 48 dihedral restraints. The atomic rms distributions about the mean coordinate for the ensemble of structures is 0.55( +/- 0.09) A for backbone atoms and 1.10( +/- 0.08) A for all heavy atoms. The protein has a topology very similar to that of telokin and the titin Ig domains and thus it falls into the I set of the immunoglobulin superfamily. The close agreement between the predicted and observed structures of Ig 18' demonstrates clearly that the I set profile can be applied in the structure prediction of immunoglobulin-like domains of diverse modular proteins. Folding studies reveal that the protein has relatively low thermodynamic stability, deltaG(H2O)U-F = 4.0 kcal mol(-1) at physiological pH. Unfolding studies suggest that the protein has considerable kinetic stability, the half life of the unfolding is greater than 40 minutes in the absence of denaturant.     

The predictors of quality care

Patients' perceptions of quality are important to the performance ratings, financial viability, and operational options afforded to purchasers of health care. Through TRICARE, the Department of Defense provides care for the more than 1.5 million active duty service members charged with the defense of the nation. As patients, these service members are typically incapable of assessing the technical quality of the care they receive. This study examines the attributes of health care delivery that define military patients' perceptions of quality and discusses the implications of these assessments.     

Purification and crystallization of complexes modeling the active state of the fragile histidine triad protein

Fragile histidine triad protein (Fhit) is a diadenosine triphosphate (ApppA) hydrolase encoded at the human chromosome 3 fragile site which is frequently disrupted in tumors. Reintroduction of FHIT coding sequences to cancer cell lines with FHIT deletions suppressed the ability of these cell lines to form tumors in nude mice even when the reintroduced FHIT gene had been mutated to allow ApppA binding but not hydrolysis. Because this suggested that the tumor suppressor activity of Fhit protein depends on substrate-dependent signaling rather than ApppA catabolism, we prepared two crystalline forms of Fhit protein that are expected to model its biologically active, substrate-bound state. Wild-type and the His96Asn forms of Fhit were overexpressed in Escherichia coli, purified to homogeneity and crystallized in the presence and absence of ApppA and an ApppA analog. Single crystals obtained by vapor diffusion against ammonium sulfate diffracted X-rays to beyond 2.75 A resolution. High quality native synchrotron X-ray data were collected for an orthorhombic and a hexagonal crystal form.      

Effect of dietary zinc levels on health and productivity of gilts and sows through two parities

The influence of 0, 50, 500 or 5,000 ppm supplemental Zn on productive characteristics, weight change, and serum and organ mineral concentrations of 60 crossbred and purebred Yorkshire gilts was evaluated. Gilts were fed their respective treatment from 30 kg body weight until the completion of two parities. Sows fed 5,000 ppm supplemental Zn weighed significantly less than sows from the other treatments when killed. Serum alkaline phosphatase activity was higher for the sows fed the highest level of Zn in all replications at 10 and 14 mo of age than for sows from the other treatments. Sows fed 0, 50 or 500 ppm had lower serum Zn and higher serum Cu concentrations than sows fed 5,000 ppm Zn at 10 and 14 mo of age. The number of pigs farrowed (total and live) and birth weight were not affected by dam's dietary treatment. However, sows receiving no additional Zn had a higher number of abnormal pigs/litter than sows on the other treatments. Sows fed 5,000 ppm additional Zn weaned fewer pigs that weighed less at weaning than sows on the other treatments. The concentration of Zn in the sow's liver increased significantly and liver Cu decreased as dietary level of Zn increased. Sows receiving 5,000 ppm Zn had lower hepatic Fe stores compared with sows receiving 500 ppm Zn. Elevated renal Cu and Zn concentrations were found in sows fed the highest level of Zn supplementation. The Zn concentration was higher and the Cu concentration lower in the aorta of sows fed 5,000 ppm Zn compared with sows fed 0 or 50 ppm additional Zn. Incidence of osteochondrosis was higher in sows supplemented with 5,000 ppm Zn than for sows from the other treatments.     

Proinflammatory macrophage-activating properties of the novel phospholipid diacylglycerol pyrophosphate

We have found that the novel phospholipid diacylglycerol pyrophosphate (DGPP), identified in bacteria, yeast, and plants, but not in mammalian cells, is able to potently activate macrophages for enhanced secretion of arachidonate metabolites, a key event in the immunoinflammatory response of leukocytes. Macrophage responses to DGPP are specific and are not mediated by its conversion into other putative lipid mediators such as phosphatidic acid, lysophosphatidic acid, or diacylglycerol. The responses to DGPP are compatible with a receptor-recognition event because they are blocked by suramin. Intracellular signaling initiated by DGPP includes phosphorylation and activation of the Group IV cytosolic phospholipase A2 and of the extracellular-signal regulated p42 mitogen-activated protein kinase (MAPK) and p44 MAPK, and membrane translocation of the protein kinase C isoenzymes alpha, epsilon, delta. These results establish DGPP as a novel macrophage-activating factor and suggest a potential role for this compound in triggering homeostatic cellular responses.     

Substrate phosphorylation in the protein kinase Cgamma knockout mouse

The phosphorylation state of three identified neural-specific protein kinase C substrates (RC3, GAP-43/B-50, and MARCKS) was monitored in hippocampal slices of mice lacking the gamma-subtype of protein kinase C and wild-type controls by quantitative immunoprecipitation following 32Pi labeling. Depolarization with potassium, activation of glutamate receptors with glutamate, or direct stimulation of protein kinase C with a phorbol ester increased RC3 phosphorylation in wild-type animals but failed to affect RC3 phosphorylation in mice lacking the gamma-subtype of protein kinase C. Our results suggests the following biochemical pathway: activation of a postsynaptic (metabotropic) glutamate receptor stimulates the gamma-subtype of protein kinase C, which in turn phosphorylates RC3. The inability to increase RC3 phosphorylation in mice lacking the gamma-subtype of protein kinase C by membrane depolarization or glutamate receptor activation may contribute to the spatial learning deficits and impaired hippocampal LTP observed in these mice.