A new method for the reconstitution of the anion transport system of the human erythrocyte membrane

Summary The anion transport protein of the human erythrocyte membrane, band 3, was solubilized and purified in solutions of the non-ionic detergent Triton X-100. It was incorporated into spherical lipid bilayers by the following procedure: (1) Dry phosphatidylcholine was suspended in the protein solution. Octylglucopyranoside was added until the milky suspension became clear. (2) The sample was dialyzed overnight against detergentfree buffer. (3) Residual Triton X-100 was removed from the opalescent vesicle suspension by sucrose density gradient centrifugation and subsequent dialysis. Sulfate efflux from the vesicles was studied, under exchange conditions, using a filtration method. Three vesicle subpopulations could be distinguished by analyzing the time course of the efflux. One was nearly impermeable to sulfate, and efflux from another was due to leaks. The largest subpopulation, however, showed transport characteristics very similar to those of the anion transport system of the intact erythrocyte membrane: transport numbers (at 30°C) close to 20 sulfate molecules per band 3 and min, an activation energy of approx. 140 kJ/mol, a pH maximum at pH 6.2, saturation of the sulfate flux at sulfate concentrations around 100mm, inhibition of the flux by H₂DIDS and flufenamate (approx.K _l-values at 30°C: 0.1 and 0.7 m, respectively), and right-side-out orientation of the transport protein (as judged from the inhibition of sulfate efflux by up to 98% by externally added H₂DIDS). Thus, the system represents, for the first time, a reconstitution of all the major properties of the sulfate transport across the erythrocyte membrane.     

Effect of hypoproteinemia on lung fluid balance in awake newborn lambs

To study the influence of plasma protein concentration on fluid balance in the newborn lung, we measured pulmonary arterial and left atrial pressures, lung lymph flow, and concentrations of protein in lymph and plasma of eight lambs, 2-3 wk old, before and after we reduced their plasma protein concentration from 5.8 +/- 0.3 to 3.6 +/- 0.6 g/dl. Each lamb underwent two studies, interrupted by a 3-day period in which we drained protein-rich systemic lymph through a thoracic duct fistula and replaced fluid losses with feedings of a protein-free solution of electrolytes and glucose. Each study consisted of a 2-h control period followed by 4 h of increased lung microvascular pressure produced by inflation of a balloon in the left atrium. Body weight and vascular pressures did not differ significantly during the two studies, but lung lymph flow increased from 2.6 +/- 0.1 ml/h during normoproteinemia to 4.1 +/- 0.1 ml/h during hypoproteinemia. During development of hypoproteinemia, the average difference in protein osmotic pressure between plasma and lymph decreased by 1.6 +/- 2 Torr at normal left atrial pressure and by 4.9 +/- 2.2 Torr at elevated left atrial pressure. When applied to the Starling equation governing microvascular fluid balance, these changes in liquid driving pressure were sufficient to account for the observed increases in lung fluid filtration; reduction of plasma protein concentration did not cause a statistically significant change in calculated filtration coefficient. Protein loss did not influence net protein clearance from the lungs nor did it accentuate the increase in lymph flow associated with left atrial pressure elevation.(ABSTRACT TRUNCATED AT 250 WORDS)     

Characterization and immunohistochemical localization of rat salivary cobalamin-binding protein and comparison with human salivary haptocorrin

Rat saliva contains a cobalamin-binding protein that binds cobalamin as well as cobinamide. The protein binds cobalamin with an affinity constant of 8 X 10(10) l X mol-1, and it binds cobalamin over a more narrow pH range (pH 7.5-10) than does human haptocorrin. It has a Stokes radius of 2.45 nm as compared to the Stokes radius of 4.50 nm for human haptocorrin. Upon isoelectricfocusing it dissociates into four strong bands with pI between 7 and 8, while human haptocorrin dissociates into acid isoproteins. Since human haptocorrin binds to concanavalin A while rat haptocorrin does not, we suggest that rat haptocorrin lacks carbohydrate. The substance concentration of rat saliva haptocorrin is 0.04-12.9 nmol X l-1 (median 7.5 nmol X l-1, n = 9) for control animals. After stimulation with isoproterenol, a beta-adrenergic agent, the substance concentration is 46.4-96.6 nmol X l-1 (median 69.7 nmol X l-1, n = 8). Immunohistochemical studies show haptocorrin in the secretory acini of the submandibular and parotid glands of the rat. In the human submandibular gland, the protein is detected both in the mucous secretory acini and in the intercalated ducts.     

Distribution of Alginate Lyase Activity among Strains of Bacillus circulans

Strains from four different DNA relatedness groups of Bacillus circulans showed apparent alginate lyase activity; the activity of three strains examined had mannuronidase specificity. A representative strain of group 4 also produced apparent inducible unsulfated chrondroitin lyase activity.     

The food of Atlantic salmon, Salmo salar L., caught by long–line in northern Norwegian waters

A total of 1145 stomachs from Atlantic salmon caught over the shelf off Helgeland/Trøndelag, and in the oceanic waters off Andenes, northern Norway during late winter–spring, 1969–1972 were examined. Food was found in 52.9% of the stomachs examined. The most important food items found in fish caught in the Helgeland/Trøndelag area were euphausids and hyperid amphipods while the myctophid Benthosema glaciale , the squid Gonatus fabricil and euphausids were found most frequently in the salmon caught off Andenes. Most salmon had preyed upon only one species, and few stomachs contained three or more prey species. The type of food did not appear to be related to the length of the fish. It is suggested that some mesopelagic feeding occurred.     

Salicylic acid blocks indomethacin-induced cyclooxygenase inhibition and lesion formation in rat gastric mucosa

Salicylic acid has been shown to decrease gastric mucosal lesions induced by indomethacin in the rat. In vitro, it has also been shown to counteract the inhibitory effect of indomethacin and aspirin on the cyclooxygenase enzyme system in seminal vesicle microsomes and in platelets and vascular tissue. The hypothesis that the mechanism of salicylic acid "protection" against indomethacin-induced gastric lesions involves interference with indomethacin-induced mucosal cyclooxygenase inhibition was tested. Male, fasted rats were treated with intragastric salicylic acid in doses of 50, 100, 200, 300, or 400 mg/kg concomitantly with a sc injection of 20 mg/kg of indomethacin. Gastric mucosal lesions and mucosal cyclooxygenase activity (as measured by ex vivo prostaglandin F2 alpha synthesis) were examined 3 hr later. Intragastric salicylic acid, 200-400 mg/kg, significantly reduced indomethacin-induced lesion formation, while counteracting significantly indomethacin inhibition of prostaglandin synthesis. Salicylic acid alone did not significantly change cyclooxygenase activity. It is concluded that topical salicylic acid can decrease indomethacin-induced gastric mucosal lesion in the rat, in part, by counteracting the inhibitory effect of indomethacin at the cyclooxygenase level.     

A high-affinity folate binding protein in normal human leukocytes: Ligand binding characteristics,ionic charge and molecular size

High-affinity binding of [³H]folate to supernatant from homogenized human leukocytes containing large amounts of binding protein displayed apparent positive cooperativity. The DEAE-Sepharose^® CL-6B chromatographic profile of the supernatant at pH 6.3 contained a major peak of folate binding (M_r approx. 25 000) in the front effluent and a smaller more acidic peak (M_r approx. 25 000) that emerged after a rise in NaCl from 30 mmol/l to 1 mol/l. Triton X-100 solubilized ceil sediment from the leukocyte homogenate contained some high-affinity folate binding activity (M_r approx 25 000), typically 5–10% of the total binding activity.     

Effect of β-FNA on Opiate δ Receptor Binding

Abstract: (β-FNA, the β -fumaramate methyl ester of naltrexone, has been shown to antagonize irreversibly the actions of morphine on the guinea pig ileum and mouse vas deferens bioassays but does not affect the actions of δ-receptor ligands on the mouse vas deferens bioassay, suggesting that the compound does not irreversibly bind to the S receptor. In this paper we examine the effect of (β -FNA on the binding of the prototypic δ agonists, Leuenkephalin and d -Ala²- d -Leu⁵-enkephalin, its metabolically stable analogue, and show that treatment of membranes with β -FNA does lead to alterations in the in vitro properties of δ receptors.