A microanalytic study of particles transport across the alveoli: role of blood platelets

Following intratracheal injection of particles of colloidal gold (30 nm) in the rat, particles in the blood platelets of the alveolar capillaries can rapidly be observed. The presence of the gold is confirmed by microanalysis. The role of this phenomenon in pulmonary clearance is discussed.     

Effects of low temperature on respiration and uptake of rubidium ions by excised barley and corn roots

The effect of temperature upon ion uptake and respiration was investigated with excised roots of corn (Zea mays) and barley (Hordeum vulgare). A strong inhibition (Q₁₀ = 5 to 8) of ion uptake was observed at temperatures below 10 C. At higher temperatures more normal temperature dependencies (Q₁₀ = 1.3 to 2) were obtained. When the data were plotted according to the Arrhenius relationship, two different activation energies were indicated above and below 10 C. Other studies have related such changes with temperature in activation energy of processes to changes in membrane properties induced by temperature. These results suggest that such phase transitions may affect ion uptake processes. If so, then differences among species in their capacity to maintain normal root function at low soil temperature and to resist low temperature stress may be related to differences in the physical properties of cellular membranes.     

Fixation of O(2) during Photorespiration: Kinetic and Steady-State Studies of the Photorespiratory Carbon Oxidation Cycle with Intact Leaves and Isolated Chloroplasts of C(3) Plants

Mass spectrometric techniques were used to trace the incorporation of [¹⁸O]oxygen into metabolites of the photorespiratory pathway. Glycolate, glycine, and serine extracted from leaves of the C₃ plants, Spinacia oleracea L., Atriplex hastata, and Helianthus annuus which had been exposed to [¹⁸O]oxygen at the CO₂ compensation point were heavily labeled with ¹⁸O. In each case one, and only one of the carboxyl oxygens was labeled. The abundance of ¹⁸O in this oxygen of glycolate reached 50 to 70% of that of the oxygen provided after only 5 to 10 seconds exposure to [¹⁸O]oxygen. Glycine and serine attained the same final enrichment after 40 and 180 seconds, respectively. This confirms that glycine and serine are synthesized from glycolate.

The labeling of photorespiratory intermediates in intact leaves reached a mean of 59% of that of the oxygen provided in the feedings. This indicates that at least 59% of the glycolate photorespired is synthesized with the fixation of molecular oxygen. This estimate is certainly conservative owing to the dilution of labeled oxygen at the site of glycolate synthesis by photosynthetic oxygen. We examined the yield of ¹⁸O in glycolate synthesized in vitro by isolated intact spinach chloroplasts in a system which permitted direct sampling of the isotopic composition of the oxygen at the site of synthesis. The isotopic enrichment of glycolate from such experiments was 90 to 95% of that of the oxygen present during the incubation.

The carboxyl oxygens of 3-phosphoglycerate also became labeled with ¹⁸O in 20- and 40-minute feedings with [¹⁸O]oxygen to intact leaves at the CO₂ compensation point. Control experiments indicated that this label was probably due to direct synthesis of 3-phosphoglycerate from glycolate during photorespiration. The mean enrichment of 3-phosphoglycerate was 14 ± 4% of that of glycine or serine, its precursors of the photorespiratory pathway, in 10 separate feeding experiments. It is argued that this constant dilution of label indicates a constant stoichiometric balance between photorespiratory and photosynthetic sources of 3-phosphoglycerate at the CO₂ compensation point.

Oxygen uptake sufficient to account for about half of the rate of ¹⁸O fixation into glycine in the intact leaves was observed with intact spinach chloroplasts. Oxygen uptake and production by intact leaves at the CO₂ compensation point indicate about 1.9 oxygen exchanged per glycolate photorespired. The fixation of molecular oxygen into glycolate plus the peroxisomal oxidation of glycolate to glyoxylate and the mitochondrial conversion of glycine to serine can account for up to 1.75 oxygen taken up per glycolate.

These studies provide new evidence which supports the current formulation of the pathway of photorespiration and its relation to photosynthetic metabolism. The experiments described also suggest new approaches using stable isotope techniques to study the rate of photorespiration and the balance between photorespiration and photosynthesis in vivo.

     

Plasma cholinesterase polymorphism in Down's syndrome.

The plasma cholinesterase variants in 80 patients having Down's syndrome have been examined and compared with a random control sample of individuals domiciled in the same area. Neither a change in frequency of known C5+ variant nor any new variant has been found in the electrophoretic studies. A highly significant increase in frequency of the fluoride resistant gene as well as a significant decrease in frequency of the atypical gene has been found in the Down's syndrome individuals. This altered distribution of the rare phenotype of plasma cholinesterase in mental illness may lead to a useful diagnostic acid in genetic counselling.     

Correlations between the thermal stability of chloroplast (thylakoid) membranes and the composition and fluidity of their polar lipids upon acclimation of the higher plant,Nerium oleander,to growth temperature

The thermal stability of excitation transfer from pigment proteins to the Photosystem II reaction center of Nerium oleander adjusts by 10 Celsius degrees when cloned plants grown at 20°C/15°C, day/night growth temperatures are shifted to 45°C/32°C growth temperature or vice versa. Concomitant with this adjustment is a decrease in the fluidity of thylakoid membrane polar lipids as determined by spin labeling. The results are consistent with the hypothesis that there is a limiting maximum fluidity compatible with maintenance of native membrane structure and function. This limiting fluidity was about the same as for a number of other species which exhibit a range of thermal stabilities. Inversely correlated shifts in lipid fluidity and thermal stability occurred during the time course of acclimation of N. oleander to new growth temperatures. Thus, the temperature at which the limiting fluidity was reached changed during acclimation while the limiting fluidity remained constant. Although the relative proportion of the major classes of membrane polar lipids remained constant during adjustments in fluidity, large changes occured in the abundance of specific fatty acids. These changes were different for the phospho- and galacto-lipids suggesting that the fatty acid composition of these two lipid classes is regulated by different mechanisms. Comparisons between membrane lipid fluidity and fatty acid composition indicate that fluidity is not a simple linear function of fatty acid composition.     

Ubiquinol-cytochrome c oxidoreductase of higher plants. Isolation and characterization of the bc1 complex from potato tuber mitochondria.

A procedure is described for isolation of active ubiquinol-cytochrome c oxidoreductase (bc1 complex) from potato tuber mitochondria using dodecyl maltoside extraction and ion exchange chromatography. The same procedure works well with mitochondria from red beet and sweet potato. The potato complex has at least 10 subunits resolvable by gel electrophoresis in the presence of dodecyl sulfate. The fifth subunit carries covalently bound heme. The two largest ("core") subunits either show heterogeneity or include a third subunit. The purified complex contains about 4 mumol of cytochrome c1, 8 mumol of cytochrome b, and 20 mumol of iron/g of protein. The complex is highly delipidated, with 1-6 mol of phospholipid and about 0.2 mol of ubiquinone/mol of cytochrome c1. Nonetheless it catalyzes electron transfer from a short chain ubiquinol analog to equine cytochrome c with a turnover number of 50-170 mol of cytochrome c reduced per mol of cytochrome c1 per s, as compared with approximately 220 in whole mitochondria. The enzymatic activity is stable for weeks at 4 degrees C in phosphate buffer and for months at -20 degrees C in 50% glycerol. The activity is inhibited by antimycin, myxothiazol, and funiculosin. The complex is more resistant to funiculosin and diuron than the beef heart enzyme. The optical difference spectra of the cytochromes were resolved by analysis of full-spectrum redox titrations. The alpha-band absorption maxima are 552 nm (cytochrome c1), 560 nm (cytochrome b-560), and 557.5 + 565.5 nm (cytochrome b-566, which has a split alpha-band). Extinction coefficients appropriate for the potato cytochromes are estimated. Despite the low lipid and ubiquinone content of the purified complex, the midpoint potentials of the cytochromes (257, 51, and -77 mV for cytochromes c1, b-560, and b-566, respectively) are not very different from values reported for whole mitochondria. EPR spectroscopy shows the presence of a Rieske-type iron sulfur center, and the absence of centers associated with succinate and NADH dehydrogenases. The complex shows characteristics associated with a Q-cycle mechanism of redox-driven proton translocation, including two pathways for reduction of b cytochromes by quinols and oxidant-induced reduction of b cytochromes in the presence of antimycin.     

Effects of stable flies (Diptera: Muscidae) and heat stress on weight gain and feed efficiency of feeder cattle.

Cattle respond to the feeding of stable flies, Stomoxys calcitrans (L.), by bunching to protect their front legs. This bunching can increase heat stress which indirectly accounts for much of the reduction in cattle weight gains. We used fly-screened, self-contained feedlot pens which allowed regulation of fly populations feeding on cattle. The indirect fly effects (bunching and heat stress) accounted for 71.5% of the reduced weight gain. The direct effect of the biting flies and energy loss involved in fighting flies accounted for 28.5% of the reduced weight gain.     

Carrier-like behaviour from a static but electrically responsive model pore.

Because of the low dielectric constant of most proteins and lipids, the electric field of an ion passing through a narrow pore is long range and will interact with neighbouring ionizable residues of the channel protein. The electrical structure of the channel may thus change transiently in response to an ion passing through the pore. Model calculations then reveal that the ratio of the unidirectional ion fluxes may approach 1 as expected for a carrier or shuttling ionophore rather than the Ussing ratio expected for a pore. Saturation behaviour also becomes carrier-like. Computer simulation is reported showing a continuous variation between pore-like and carrier-like behaviour as the parameters of the system are allowed to change smoothly.     

A hybrid protein of urokinase growth-factor domain and plasminogen-activator inhibitor type 2 inhibits urokinase activity and binds to the urokinase receptor.

The binding of urokinase-type plasminogen activator (uPA) to its specific cell-surface receptor (uPAR) localises the proteolytic cascade initiated by uPA to the pericellular environment. Inhibition of uPA activity or prevention of uPA binding to uPAR might have a beneficial effect on disease states wherein this activity is deregulated, e.g. cancer and some inflammatory diseases. To this end, a bifunctional hybrid molecule consisting of the uPAR-binding growth-factor domain of uPA (amino acids 1-47; GFuPA) at the N-terminus of plasminogen-activator inhibitor type 2 (PAI-2) was produced in Saccharomyces cerevisiae. The purified protein inhibited uPA with kinetics similar to placental or recombinant PAI-2 and was also found to bind to U937 cells and to FL amnion cells. GFuPA-PAI-2 competed with uPA, the N-terminal fragment of uPA and a proteolytic fragment of uPA (amino acids 4-43) in cell binding experiments, indicating that the molecule bound to the cells via uPAR. Hence, both the uPA-inhibitory and uPAR-binding domains of the hybrid molecule were functional, demonstrating the feasibility of the novel concept of introducing an unrelated, functional domain onto a member of the serine-protease-inhibitor superfamily.