Conformational transition between native and reactive center cleaved forms of alpha 1-antitrypsin by Fourier transform infrared spectroscopy and small-angle neutron scattering

alpha 1-Antitrypsin (alpha 1-AT) is the best-characterized member of the serpin superfamily of plasma proteins. Protease inhibitor members of this family undergo a characteristic reactive-center cleavage during expression of their inhibitory activity. The physical basis of this transition in alpha 1-AT from the stressed native conformation to the more stable reactive center cleaved (split) form was studied by Fourier transform infrared (FT-IR) spectroscopy and neutron scattering. The FT-IR spectra show that, while split alpha 1-AT has three intense well-resolved components associated with the presence of antiparallel beta-sheet and alpha-helix conformations, the amide I band of native alpha 1-AT has only one intense component, associated with the presence of beta-sheet structure. 1H-2H exchange within the polypeptide backbone, studied by FT-IR and NMR spectroscopy, shows that the native form undergoes greater exchange than the split form. Under the same conditions, neutron scattering shows no differences in the radius of gyration RG of the native and the split forms. In contrast, in high concentrations of phosphate approaching those used for crystallization, the native form (unlike the split form) undergoes dimerization. These data indicate that the conformational transition largely involves localized secondary and tertiary structure rearrangements. We propose that the energetically stressed native alpha 1-AT structure is the consequence of a significantly reduced number of hydrogen bonds in secondary structure components and that reactive-site cleavage between Met358 and Ser359 is the key for the development of the fully hydrogen bonded more stable serpin structure.     

Synthesis and degradation of fatty acid ethyl esters by cultured hepatoma cells exposed to ethanol

Fatty acid ethyl esters are a family of neutral lipids that are the products of esterification of fatty acids with ethanol. Unlike other pathways of ethanol metabolism, ethyl esters are present in numerous human organs which are the targets of ethanol-induced damage. In the present study, we have shown that fatty acid ethyl esters are synthesized by a hepatoma cell line in tissue culture when exposed to ethanol concentrations easily attained by man during social drinking. Unlike alcohol dehydrogenase, the enzyme(s) responsible for synthesis of ethyl esters are membrane-bound and concentrated in the microsomal fraction of rat hepatocytes. In addition, fatty acid ethyl esters are hydrolyzed to free fatty acids and ethanol by membrane-bound enzyme(s) that are enriched in the microsomal and mitochondrial-lysosomal fractions. Intracellular hydrolysis of fatty acid ethyl esters release free fatty acids which are preferentially incorporated into cellular cholesterol esters. Thus, we have shown that a hepatocellular line exposed to concentrations of ethanol easily achieved in man by social drinking utilize endogenous fatty acids to form long-lived ethanol metabolites, fatty acid ethyl esters. Importantly, this family of neutral lipids may act as biochemical mediators of ethanol-induced cell damage, including the changes in cholesterol metabolism noted in chronic alcoholics.     

Two-Dimensional Electrophoretic Mapping of Proteins of Bundle Sheath and Mesophyll Cells of the C(4) Grass Digitaria sanguinalis (L.) Scop. (Crabgrass)

Two-dimensional electrophoresis was performed on proteins of bundle sheath and mesophyll cells isolated from the C₄ grass Digitaria sanguinalis (L.) Scop. Two-dimensional maps of these proteins were constructed and ribulose-1,5-biphosphate carboxylase and phosphoenolpyruvate carboxylase were identified. Of the total number of proteins found in both cell types, 36% were found only in bundle sheath cells, 17% only in mesophyll cells, and 47% in both cell types. By comparison, the distributions of 48 enzymes assayed in these cell types were 35%, 21%, and 44%, respectively.

Protein patterns were also compared with C₄ plants exhibiting different decarboxylation pathways and, in both bundle sheath and mesophyll cells, proteins were found which were unique to each species. Bundle sheath proteins of one C₄ species were found to be more like bundle sheath proteins of another C₄ species than like mesophyll proteins of the same species.

     

Habitat assortment of sexes and water balance in a dioecious grass

Summary Eggs of the carabid beetle Pterostichus oblongopunctatus kept under optimum conditions in petri dishes had a mean mortality of 8% (n=200), while the mortality of eggs incubated in fresh litter was 83% (n=250). Heating the litter up to 70° C and thus eliminating other organisms from the litter fauna reduced mortality to 18% (n=250). When eggs were deposited by the females themselves in a perspex cylinder containing an undisturbed soil sample, the mean egg mortality was 70% (n235), but when predators had been eliminated the mortality was only 7% (n235). The assumption of Grüm (1975) that egg mortality can be neglected does not hold for P. oblongopunctatus. The predators of the eggs must be common species, but it is not yet known to which family they belong.     

Glucocorticoid binding to solubilized components of human placental trophoblast membranes

We have recently described glucocorticoid uptake by human placental membrane vesicles which is specific, saturable and has a low K (7 nM). This paper describes solubilization of these vesicles with Triton x-100 and successful demonstration of glucocorticoid binding to the putative transport site. This was accomplished by analysis of corticosterone binding to the 140,000 × g supernatant of solubilized vesicles using G-75 Sephadex chromatography. The amount of bound corticosterone present in the void volume was proportional to the concentration of solubilized vesicles. The specificity of binding was tested by coincubation of tritiated corticosterone with 100-fold excesses of various steroids. The relative ability of various steroids to inhibit binding was corticosterone=pregesterone- >aldosterone. Triamcinolone acetonide, and estradiol were ineffective competitors. We conclude from these studies that human placental membranes contain glucocorticoid-specific binding sites which can be solubilized with Triton x-100. It is possible that these sites are involved in membrane mediated transport of glucocorticoids by this tissue.     

Persistence of predator-prey systems in an uncertain environment

Summary The time derivatives of prey and predator populations are assumed to satisfy a set of inequalities, instead of a precise differential equation, reflecting an uncertain environmental and/or lack of knowledge by the modeler. A system of differential equations is found whose solution gives the boundary of a persistent set, which is positive flow invariant for any system satisfying the inequalities. Conditions are given for the persistent set to be bounded away from both axes, which show that resonance effects cannot drive either predator or prey to extinction if that does not happen for an autonomous system satisfying the inequalities. In general predator-prey systems are more persistent when there is strong asymptotic stability, when there is correlation between prey and predator dynamics, when the effect of perturbations is density dependent, and are more persistent under perturbations of the prey than of the predator.     

Scanning electron microscopy of taxonomic diagnostic criteria of the freshwater sponge,Heteromeyenia tubisperma (Potts, 1881) (Porifera : Spongillidae)

Taxonomic diagnostic criteria of the spongillid freshwater sponge, Heteromeyenia tubisperma (Potts, 1881) were examined using scanning electron microscopy. The species is characterized by a gemmule which bears an unusually long, prominent porous tube. The application of SEM to the systematic studies of the freshwater sponges provides diagnostic capabilities not available with the light microscope. It is desirable that a key, coupled with a reference atlas of scanning electron micrographs illustrating taxonomic diagnostic criteria of freshwater sponge species, particularly utilizing type specimens, be developed.     

Survival of Diplectrona modesta banks (Trichoptera : Hydropsychidae) during short periods of desiccation

Larvae of the caddisfly, Diplectrona modesta (Hydropsychidae), were found to be permanent inhabitants of a small, intermittent, headwater stream in southern Ontario. They survived desiccation for at least four weeks in summer. A laboratory experiment gave a mean survivorship of 50% after 30 days without water in stream gravel and stones after the larvae had been permitted to construct their shelters. Survivors were those that constructed their tubes deep in the substrate.     

Cationic responses of organs and haemolymph of Biomphalaria pfeifferi (Krauss), Biomphalaria glabrata (Say) and Helisoma trivolvis (Say) (Gastropoda: Planorbirdae) to cationic alterations of the medium

The cationic responses of haemolymph, mantle collar, headfoot, gut + digestive diverticulum and ovotestis + albumin gland of three planorbids to cationic changes in the media are explored. Body organs exhibited cationic homeostasis, although attained with difficulty under very low Ca : Mg and Ca : Na ratios. Haemolymph imbalances were obtained at very low calcium concentrations and at very low Ca : Mg and Ca : Na ratios in the medium; this may be linked to competition for Ca²⁺ uptake sites in the epithelium by Mg²⁺ and possibly Na⁺ ions.Fecundity and shell growth in relation to calcium concentrations and to Ca : Mg and Ca : Na ratios are examined.Normal internal cationic levels, under optimal conditions, were obtained for each species. Significant interspecific differences for haemolymph sodium were found; magnesium levels were slightly higher in Biomphalaria spp. than in Helisoma trivolvis; potassium levels were all similar. Amounts of tissue calcium decreased as follows: Mantle collar head-foot ovotestis + albumin gland-gut + diverticulum. Tissue magnesium levels in the gut were low. H. trivolvis had highest tissue calcium and sodium; B. glabrata had highest tissue potassium.