Prediction of sequential antigenic regions in proteins

Prediction of antigenic regions in a protein will be helpful for a rational approach to the synthesis of peptides which may elicit antibodies reactive with the intact protein. Earlier methods are based on the assumption that antigenic regions are primarily hydrophilic regions at the surface of the protein molecule. The method presented here is based on the amino acid composition of known antigenic regions in 20 proteins which is compared with that of 314 proteins [(1978) Atlas of Protein Sequence and Structure, vol. 5, suppl. 3, 363-373]. Antigenicity values were derived from the differences between the two data sets. The method was applied to bovine ribonuclease, the B-subunit of cholera toxin and herpes simplex virus type 1 glycoprotein D. There was a good correlation between the predicted regions and previously determined antigenic regions.     

An experimental analysis of cache recovery in Clark's nutcracker

Five hypotheses of cache recovery behaviour in Clark's nutcracker (Nucifraga columbiana) were examined experimentally. Most caches were made in soil within 5 cm of conspicuous large objects. Both seed-caching and non-seed-caching nutcrackers were able to locate caches. Seed-caching nutcrackers relocated caches using large objects as remembered visual cues. Soil microtopography and small (<2 cm diameter) objects may be used as cues to facilitate cache recovery but are not essential. Non-seed-caching nutcrackers located caches by using soil disturbances at cache sites as visual cues and by searching preferentially near objects where caches were concentrated. Success rates of seed-caching nutcrackers ranged from 52 to 78% and those of non-seed-caching nutcrackers ranged from 8 to 12%. Nutcrackers do not use random search or olfactory cues to locate caches.     

Analogs of oxytocin containing a modified peptide bond

Analogs of deamino-oxytocin and deamino-oxypressin containing a CH2-NH group instead of an amide bond between positions 8 and 9 were synthesized. All tested compounds exhibit significantly lowered biological activities.     

Increased demand for NAD+ relative to ATP drives aerobic glycolysis

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Insulin-mediated modifications of myocardial lipoprotein lipase and lipoprotein metabolism

Recirculating organ perfusion in vitro was conducted with hearts from control rats, animals given a single dose of streptozotocin (65 mg/kg) 48 h earlier, and streptozotocin-treated rats administered insulin (5 units), 2 h prior to organ perfusion. During 45-min perfusions, the lipolysis of very low density lipoprotein (VLDL) triglyceride was significantly less in hearts from diabetics than in controls (41.9 +/- 7.3% of control). This was associated with significant reductions in heparin-releasable (functional) lipoprotein lipase and tissue lipoprotein lipase of perfused hearts. The decreases in VLDL triglyceride metabolism and the levels of myocardial lipoprotein lipase were completely reversed by treatment of diabetic rats with insulin 2 h prior to study. Similar improvement of VLDL triglyceride metabolism and increases in myocardial lipoprotein lipase activity were observed in hearts from diabetic rats by direct addition of 100 milliunits/ml of insulin to the recirculating perfusion media. Under these conditions, the increase in both fractions of lipoprotein lipase in response to insulin was completely inhibited, and utilization of VLDL triglyceride was partially inhibited by pre-perfusion with cycloheximide for 10 min. The data derived from either VLDL triglyceride lipolysis in organ perfusion or direct measurement of myocardial lipoprotein lipase demonstrate a direct effect of insulin on myocardial lipoprotein lipase activity, and suggest that the response to insulin may be due in part to effects on protein synthesis.     

The oxidation of the calcium probe quin2 and its analogs by prostaglandin H synthase

Quin2 and its analogs BAPTA, 5,5'-dimethyl BAPTA, 5,5'-difluoro BAPTA, fura-2, and indo-1 were developed to measure intracellular calcium concentrations. In this study we investigated whether quin2 and its analogs are susceptible to peroxidase-catalyzed oxidation. The hydroperoxidase activity of prostaglandin H synthase, like other peroxidases, is capable of oxidizing a wide variety of substrates. It was found that quin2 and its analogs served as reducing cofactors for the hydroperoxidase activity of prostaglandin H synthase, undergoing oxidation in the process. Furthermore, arachidonic acid metabolism was stimulated. Oxidation of quin2 and its analogs resulted in the formation of a carbon-centered radical, as could be detected by ESR, and in the formation of formaldehyde. Quin2 fluorescence decreased upon addition of arachidonic acid and prostaglandin H synthase. Furthermore, addition of calcium no longer resulted in an increase in quin2 fluorescence, as was observed prior to the addition of arachidonic acid and the enzyme. This indicates that one or more of the -N-CH2-COOH groups, which are responsible for the binding of calcium, were oxidized by the hydroperoxidase. Since prostaglandin H synthase is present in many cellular systems in which calcium concentrations are modulated, oxidation of the calcium probe might not only affect the measurement of intracellular calcium but could activate arachidonic acid metabolism as well.     

Characterization of adenine phosphoribosyltransferase from the human malaria parasite, Plasmodium falciparum

Because of their inability to synthesize purines de novo, malaria parasites rely on purine phosphoribosyltransferases (PRTases) to convert purine bases salvaged from the host cell (the erythrocyte) into the corresponding purine nucleoside monophosphates. Our studies with late trophozoites of the human malaria parasite, Plasmodium falciparum, showed that virtually all of the purine PRTase activity is accounted for by two distinct enzymes. One enzyme utilizes hypoxanthine, guanine and xanthine (Queen, S.A., Vander Jagt, D. and Reyes, P. (1988) Mol. Biochem. Parasitol. 30, 123-134). The second enzyme utilizes only adenine and is the subject of this paper. This latter enzyme exhibits a biphasic pH-activity profile and is moderately to weakly inhibited by several divalent metal ions. Several of the properties of the P. falciparum enzyme were found to differ significantly from those of human erythrocyte adenine PRTase. (1) The molecular weight (18,000) of the parasite enzyme is smaller than that of the host cell enzyme. (2) The parasite enzyme, unlike the erythrocyte enzyme, is not significantly inhibited by sulfhydryl reagents. (3) 6-Mercaptopurine and 2,6-diaminopurine proved to be competitive inhibitors of the parasite enzyme (Ki 0.70 and 1.0 mM, respectively); on the other hand, the human enzyme is not inhibited by these agents. (4) The Km for adenine (0.80 microM) and 5-phosphoribosyl-1-pyrophosphate (0.70 microM) displayed by the parasite enzyme are significantly smaller than the corresponding Km values shown by the erythrocyte enzyme. These distinctions between the parasite and host enzymes point to the possibility that adenine PRTase of P. falciparum may represent a potential target for chemotherapeutic attack.     

The influence of porphyrins on iron-catalysed generation of hydroxyl radicals.

Uroporphyrin I, haematoporphyrin and haematoporphyrin derivative had no effect on O2-. generation during oxidation of hypoxanthine by xanthine oxidase and on the formation of hydroxyl radicals (OH.) in the hypoxanthine/xanthine oxidase/Fe3+-EDTA/deoxyribose system. On the other hand, these porphyrins strongly inhibited O2-. formation in a horseradish peroxidase/H2O2/NADPH mixture, whereas they augmented OH. generation in this system after addition of Fe3+-EDTA. Experimental evidence suggests that these observations should be ascribed to the formation of a porphyrin anion radical in the horseradish peroxidase/NADPH system. The formation of this anion radical was confirmed by e.s.r. spectroscopy. This radical is apparently unable to reduce cytochrome c, but it can replace O2-. in the OH.-generating Haber-Weiss reaction.     

N2-Succinylornithine in ornithine catabolism of Pseudomonas aeruginosa

Most Pseudomonas aeruginosa PAO mutants which were unable to utilize l-arginine as the sole carbon and nitrogen source (aru mutants) under aerobic conditions were also affected in l-ornithine utilization. These aru mutants were impaired in one or several enzymes involved in the conversion of N²-succinylornithine to glutamate and succinate, indicating that the latter steps of the arginine succinyltransferase pathway can be used for ornithine catabolism. Addition of aminooxyacetate, an inhibitor of the N²-succinylornithine 5-aminotransferase, to resting cells of P. aeruginosa in ornithine medium led to the accumulation of N²-succinylornithine. In crude extracts of P. aeruginosa an ornithine succinyltransferase (l-ornithine:succinyl-CoA N²-succinyltransferase) activity could be detected. An aru mutant having reduced arginine succinyltransferase activity also had correspondingly low levels of ornithine succinyltransferase. Thus, in P. aeruginosa, these two activities might be due to the same enzyme, which initiates aerobic arginine and ornithine catabolism.Abbreviations OAT ornithine 5-aminotransferase - SOAT N²-succinylornithine 5-aminotransferase - Oru ornithine utilization - Aru arginine utilization     

Enhancement of allergic lung sensitization in mice by ozone inhalation

Inhaled ozone was found to exert an enhancing effect for allergic lung sensitization when mice contracted an aerosolized allergen. The animals were exposed to ozone concentrations of 0.24, 0.16, 0.13, and 0.10 ppm. After 4 days of continuous ozone exposure, the mice had allergen contact from an aerosolized solution of ovalbumin. The animals were then maintained in ambient air for several days before the cycle of ozone and aerosolized allergen was repeated over four allergen contact cycles. Mice were rested in ambient air for a week after the last allergen contact, and they were then tested for allergic sensitization by the intravenous injection of 2 mg of ovalbumin to induce anaphylactic shock in allergic individuals. The control groups of mice were maintained in ambient air throughout the experiment, but they experienced identical allergen contact with the ozone-exposed mice. The phenomenon of allergic enhancement from ozone inhalation was detected at 0.24, 0.16, and 0.13 ppm of ozone. The enhancing effect disappeared at 0.10 ppm of ozone. The study indicated a potential for increasing the number of allergically sensitized individuals when various allergens are inhaled during periods of high ozone exposure with the consequent adverse changes on respiratory membranes. The significance to human health of the allergic enhancement phenomenon by ozone needs investigation.