NADH oxidation and NAD+ reduction catalysed by tightly coupled inside-out vesicles from Paracoccus denitrificans.

Tightly coupled inside-out vesicles were prepared from Paracoccus denitrificans cells (SPP, sub-Paracoccus particles) and characterized kinetically. The rate of NADH oxidation, catalysed by SPP, increases 6-8 times on addition of gramicidin. The vesicles are capable of catalysing Delta micro H+-dependent reverse electron transfer from quinol to NAD+. The kinetic parameters of the NADH-oxidase and the reverse electron transfer carried out by membrane-bound P. denitrificans complex I were estimated and compared with those of the mitochondrial enzyme. The data demonstrate that catalytic properties of the dinucleotide-binding site of the bacterial and mitochondrial complex I are almost identical, pointing out similar organization of the site in mammals and P. denitrificans. Inhibition of the bacterial complex I by a specific inhibitor of Q reduction, rotenone, is very different from that of the mitochondrial enzyme. The inhibitor is capable of suppressing the NADH oxidation reaction only at micromolar concentrations, while the activity of mitochondrial enzyme is suppressed by nanomolar concentrations of rotenone. In contrast to the mitochondrial enzyme, rotenone, even at concentrations as high as 10 micro m, does not inhibit the reverse, Delta micro H+-dependent NAD+-reductase reaction on SPP.     

Explanations for the acclimation period preceding the mineralization of organic chemicals in aquatic environments

A study was conducted of possible reasons for acclimation of microbial communities to the mineralization of organic compounds in lake water and sewage. The acclimation period for the mineralization of 2 ng of p-nitrophenol (PNP) or 2,4-dichlorophenoxyacetic acid per ml of sewage was eliminated when the sewage was incubated for 9 or 16 days, respectively, with no added substrate. The acclimation period for the mineralization of 2 ng but not 200 ng or 2 micrograms of PNP per ml was eliminated when the compound was added to lake water that had been first incubated in the laboratory. Mineralization of PNP by Flavobacterium sp. was detected within 7 h at concentrations of 20 ng/ml to 2 micrograms/ml but only after 25 h at 2 ng/ml. PNP-utilizing organisms began to multiply logarithmically after 1 day in lake water amended with 2 micrograms of PNP per ml, but substrate disappearance was only detected at 8 days, at which time the numbers were approaching 10(5) cells per ml. The addition of inorganic nutrients reduced the length of the acclimation period from 6 to 3 days in sewage and from 6 days to 1 day in lake water. The prior degradation of natural organic materials in the sewage and lake water had no effect on the acclimation period for the mineralization of PNP, and naturally occurring inhibitors that might delay the mineralization were not present. The length of the acclimation phase for the mineralization of 2 ng of PNP per ml was shortened when the protozoa in sewage were suppressed by eucaryotic inhibitors, but it was unaffected or increased if the inhibitors were added to lake water.(ABSTRACT TRUNCATED AT 250 WORDS)     

Kinetics of p-nitrophenol mineralization by a Pseudomonas sp.: effects of second substrates

The kinetics of simultaneous mineralization of p-nitrophenol (PNP) and glucose by Pseudomonas sp. were evaluated by nonlinear regression analysis. Pseudomonas sp. did not mineralize PNP at a concentration of 10 ng/ml but metabolized it at concentrations of 50 ng/ml or higher. The Ks value for PNP mineralization by Pseudomonas sp. was 1.1 micrograms/ml, whereas the Ks values for phenol and glucose mineralization were 0.10 and 0.25 micrograms/ml, respectively. The addition of glucose to the media did not enable Pseudomonas sp. to mineralize 10 ng of PNP per ml but did enhance the degradation of higher concentrations of PNP. This enhanced degradation resulted from the simultaneous use of glucose and PNP and the increased rate of growth of Pseudomonas sp. on glucose. The Monod equation and a dual-substrate model fit these data equally well. The dual-substrate model was used to analyze the data because the theoretical assumptions of the Monod equation were not met. Phenol inhibited PNP mineralization and changed the kinetics of PNP mineralization so that the pattern appeared to reflect growth, when in fact growth was not occurring. Thus, the fitting of models to substrate depletion curves may lead to erroneous interpretations of data if the effects of second substrates on population dynamics are not considered.     

cDNA and protein sequence of a major pea seed albumin (PA 2 : Mr≈26 000)

Summary Pea albumin 2 (PA2:M_r26000) is a major component of the albumin fraction derived from aqueous salt extracts of pea seed. Sodium dodecylsulfate-polyacrylamide gel electrophoresis and chromatography on DEAE-Sephacel resolve PA2 into two closely related components (PA2a and PA2b). A cDNA clone coding for one of these components has been sequenced and the deduced amino acid sequence compared with partial, chemically-determined sequences for cyanogen bromide peptides from both PA2 components. Complete amino acid sequences were obtained for the C-terminal peptides. The PA2 molecule of 230 amino acids contains four imperfect repeat sequences each of approximately 57 amino acids in length.The combined sequence data, together with a comparison of PA2-related polypeptides produced in vitro and in vivo, indicate that PA2 is synthesized without a signal sequence and does not undergo significant post-translational modification. Although both forms of PA2 contain Asn-X-Thr consensus sequences, neither form is glycosylated. Accumulation of PA2 contributes approximately 11% of the sulfur-amino acids in pea seeds (cysteine plus methionine equals 2.6 residues percent). Suppression of levels of PA2 polypeptides and their mRNAs in developing seeds of sulfur-deficient plants is less marked than that for legumin, in spite of the lower content of sulfur-amino acids in legumin.     

Does carnitine have a role in fat absorption?

The effect of D-carnitine and tetradecylglycidic acid (TDGA), an inhibitor of carnitine palmitoyltransferase, on intestinal absorption of palmitic acid was determined. The proximal intestinal segment was ligated in adult male rats and filled with 0.5 microCi of 14C-palmitic acid alone or with either D-carnitine or TDGA. Thirty minutes later the radioactivity was determined in the intestinal lumen, intestinal wall and plasma. The absorption of palmitic acid was decreased in the presence of D-carnitine (10 mg/ml) as evidenced by significantly lower levels of radioactivity in the gut wall and the plasma and by significantly greater residual radioactivity in the lumenal contents. L-carnitine had no effect on plasma radioactivity but if D- and L-carnitine were given together the effect of D-carnitine was still in evidence. TDGA also inhibited intestinal absorption of palmitic acid.     

Calculation of leaf photosynthetic parameters from light-response curves for ecophysiological applications

Measurement of the light response of photosynthetic CO₂ uptake is often used as an implement in ecophysiological studies. A method is described to calculate photosynthetic parameters, such as the maximum rate of whole electron transport and dissimilative respiration in the light, from the light response of CO₂ uptake. Examples of the light-response curves of flag leaves and ears of wheat (Triticum aestivum cv. ARKAS) are shown.Abbreviations and symbols A net photosynthesis rate - D ¹ rate of dissimilative respiration occurring in the light - f loss factor - I incident PPFD - I effective absorbed PPFD - J rate of whole electron transport - J _m maximum rate of whole electron transport - p ^c intercellular CO₂ partial pressure - PPFD photosynthetic photon flux density - q effectivity factor for the use of light (electrons/quanta) - absorption coefficient - I ^* CO₂ compensation point in the absence of dissimilative respiration (bar) - II conversion factor for calculation of CO₂ uptake from the rate of whole electron transport - convexity factor Gas-exchange rates relate to the projective area and are given in mol·m^-2·s^-1. Electron-transport rates are given in mol electrons·m^-2·s^-1; PPFD is given in mol quanta·m^-2·s^-1.     

Identification of somatic hybrids in plant protoplast fusions with monoclonal antibodies to plasma-membrane antigens

Monoclonal antibodies generated by immunization with a plasma-membrane preparation from suspension-cultured cells of Nicotiana glutinosa L. were used in combination with fluoresceinor rhodamine-labeled goat anti-mouse immunoglobulins to identify heterokaryons in protoplast fusion procedures. Antibody labeling did not inhibit callus formation nor plantlet regeneration. The antibodies are non-invasive and surface labeling provides clear optical discrimination of true heterokaryons from unfused aggregates as well as from parental protoplasts and homokaryons. Labeling is stable throughout fusion and hence by pre-labeling parental protoplast populations the strategy is both versatile and of general applicability.     

Role of dissolution rate and solubility in biodegradation of aromatic compounds.

Strains of Moraxella sp., Pseudomonas sp., and Flavobacterium sp. able to grow on biphenyl were isolated from sewage. The bacteria produced 2.3 to 4.5 g of protein per mol of biphenyl carbon, and similar protein yields were obtained when the isolates were grown on succinate. Mineralization of biphenyl was exponential during the phase of exponential growth of Moraxella sp. and Pseudomonas sp. In biphenyl-supplemented media, Flavobacterium sp. had one exponential phase of growth apparently at the expense of contaminating dissolved carbon in the solution and a second exponential phase during which it mineralized the hydrocarbon. Phase-contrast microscopy did not show significant numbers of cells of these three species on the surface of the solid substrate as it underwent decomposition. Pseudomonas sp. did not form products that affected the solubility of biphenyl, although its excretions did increase the dissolution rate. It was calculated that Pseudomonas sp. consumed 29 nmol of biphenyl per ml in the 1 h after the end of the exponential phase of growth, but 32 nmol of substrate per ml went into solution in that period when the growth rate had declined. In a medium with anthracene as the sole added carbon source, Flavobacterium sp. converted 90% of the substrate to water-soluble products, and a slow mineralization was detected when the cell numbers were not increasing. Flavobacterium sp. and Beijerinckia sp. initially grew exponentially and then arithmetically in media with phenanthrene as the sole carbon source. Calculations based on the growth rates of these bacteria and the rates of dissolution of phenanthrene suggest that the dissolution rate of the hydrocarbon may limit the rate of its biodegradation.     

Serotonin induced actin polymerization and association with cytoskeletal elements in cultured bovine aortic endothelium

Serotonin, (5-hydroxytryptamine, 5-HT) binds to cultured endothelial cell stress fibers as identified by fluorescence microscopy and in vitro induces actin polymerization as measured by DNase 1 inhibition and differential centrifugation; the structural change in actin in the presence of 5-HT resembles actin paracrystals as visualized by negative staining under electron microscopy. These observations indicate that 5-HT, which is taken up by endothelium by a non-mediated diffusion, may interact directly with actin to affect cytoskeletal changes.