Biomedical subjects

R Fall

Publications and source records attributed to R Fall.

At least 37 records · Page 2Linked to original sources

PubMed1995

Characterization of aspen isoprene synthase, an enzyme responsible for leaf isoprene emission to the atmosphere.

Isoprene (2-methyl-1,3-butadiene) is a volatile hydrocarbon emitted from many plant species to the atmosphere, where it plays an important role in atmospheric chemistry. An enzyme extracted from aspen (Populus tremuloides) leaves was previously found to catalyze the Mg(2+)-dependent elimination of pyrophosphate from dimethylallyl diphosphate (DMAPP) to form isoprene (Silver, G. M., and Fall, R. (1991) Plant Physiol. 97, 1588-1591). This enzyme, isoprene synthase, has now been purified 4000-fold to near homogeneity. The enzyme had a native molecular mass of 98-137 kDa and isoelectric point of 4.7 and contained 58- and 62-kDa subunits, implying that it is a heterodimer. Partial amino acid sequences of the two subunits indicated they are closely related to each other and that they do not share a strong homology with any other reported proteins. The isoprene synthase reaction was dependent on Mg2+ or Mn2+, and the reaction products were shown to be isoprene and pyrophosphate with a stoichiometry close to 1:1. The Km for DMAPP was high at 8 mM, and the kcat of 1.7 s-1 was low, but similar to those of other allylic diphosphate-utilizing enzymes. It is argued that the isoprene synthase reaction may be much more efficient in vivo, where it is under light-dependent control. It seems probable that this unique enzyme, rather than non-enzymatic reactions, can account for the emission of hundreds of millions of metric tons of isoprene from plants to the global atmosphere each year.

Alkyl and Aryl Transferases↗

PubMed1995

Bacteria produce the volatile hydrocarbon isoprene.

Various bacterial species, both Gram-negative and Gram-positive, were found to produce the volatile hydrocarbon isoprene (2-methyl-1,3-butadiene). Out of the tested cultures, Bacillus produced the most isoprene. The production of isoprene from bacteria was confirmed by gas chromatography-mass spectrometry. Media and growth effects on isoprene production were investigated: growth in rich media led to higher levels of isoprene than growth in minimal media, and highest isoprene emission rates were seen in log-phase cultures. Temperature profiles for bacterial isoprene production showed an optimum of 45 degrees C and were suggestive of an enzymatic mechanism for isoprene formation.

Bacillus↗

PubMed1995

Marine Vibrio species produce the volatile organic compound acetone.

While screening aerobic, heterotrophic marine bacteria for production of volatile organic compounds, we found that a group of isolates produced substantial amounts of acetone. Acetone production was confirmed by gas chromatography, gas chromatography-mass spectrometry, and high-performance liquid chromatography. The major acetone producers were identified as nonclinical Vibrio species. Acetone production was maximal in the stationary phase of growth and was stimulated by addition of l-leucine but not the other common amino acids, suggesting that leucine degradation leads to acetone formation. Acetone production by marine vibrios may contribute to the dissolved organic carbon associated with phytoplankton, and some of the acetone produced may be volatilized to the atmosphere.

Journal Article↗

PubMed1994

[Control of reproduction in the female Ndama cow by Norgestomet (CRESTAR)].

The purpose of this experiment was to test the efficiency of the Norgestomet CRESTARND on 91 Ndama cows living in three different ecological zones. The results of the experiments showed an average rate of heat synchronisation of 97.8%, an average heat time of 10.17 +/- 2.81 h with the intensity of these heat being essentially low or medium levels. The heats also occurred of mostly during the night. The time lag between the PGF2 alpha injection and the first signs of oestrus was 83.96 +/- 14.96 h and the one between removing the implant and the first heats was 34.78 +/- 14.9 h and the average blood's level of progesterone was 5 +/- 10.3 ng/ml. The efficiency of CRESTARND in the control of the sexual cycle of the Ndama cows was demonstrated by this study.

Animals↗

PubMed1994

[Embryo transfer in a Senegal village environment].

This study aims to identify the embryo transfer constraints and production costs at a village level in the district of Kolda (Senegal). Fifteen (15) donors cows were superovulated using 2500 UI PMSG (group A n = 8), 32 mg of FSH (group B n = 3) and 36 mg of FSH (group C n = 4). The average number of yellow bodies that were palpated was 5.06 and the mean rates of collected embryos were 5.66 for group A, 2.5 for group B and 3.3 for group C. The average transferable embryos were 2.33 for group A, 0 for group B and 1.4 for group C. The identified constraints are nutritional, healthy, social and logistic nature. The average cost of a produced and transferred embryo amount respectively to 75.940 F CFA and 99310 F CFA. Embryo transfer can fit in with a dairy production development plan in Senegal.

Animals↗

PubMed1993

High-level expression of ice nuclei in a Pseudomonas syringae strain is induced by nutrient limitation and low temperature.

Attempts were made to maximize the expression of ice nuclei in Pseudomonas syringae T1 isolated from a tomato leaf. Nutritional starvation for nitrogen, phosphorous, sulfur, or iron but not carbon at 32 degrees C, coupled to a shift to 14 to 18 degrees C, led to the rapid induction of type 1 ice nuclei (i.e., ice nuclei active at temperatures warmer than -5 degrees C). Induction was most pronounced in stationary-phase cells that were grown with sorbitol as the carbon source and cooled rapidly, and under optimal conditions, the expression of type 1 ice nuclei increased from < 1 per 10(7) cells (i.e., not detectable) to 1 in every cell in 2 to 3 h. The induction was blocked by protein and RNA synthesis inhibitors, indicative of new gene expression. Pulse-labeling of nongrowing cultures with [35S]methionine after a shift to a low temperature demonstrated that the synthesis of a new set of "low-temperature" proteins was induced. Induced ice nuclei were stable at a low temperature, with no loss in activity at 4 degrees C after 8 days, but after a shift back to 32 degrees C, type 1 ice nuclei completely disappeared, with a half-life of approximately 1 h. Repeated cycles of low-temperature induction and high-temperature turnover of these ice nuclei could be demonstrated with the same nongrowing cells. Not all P. syringae strains from tomato or other plants were fully induced under the same culture conditions as strain T1, but all showed increased expression of type 1 ice nuclei after the shift to the low temperature. In support of this view, analysis of the published DNA sequence preceding the translational start site of the inaZ gene (R. L. Green and G. Warren, Nature [London] 317:645-648, 1985) suggests the presence of a gearbox-type promoter (M. Vincente, S. R. Kushner, T. Garrido, and M. Aldea, Mol. Microbiol. 5:2085-2091, 1991).

Bacterial Outer Membrane Proteins↗

PubMed1993

Kinetics of appearance and disappearance of classes of bacterial ice nuclei support an aggregation model for ice nucleus assembly.

The kinetics of appearance and disappearance of three classes of ice nuclei in Pseudomonas syringae was investigated under conditions where high-level expression of the ice nucleation phenotype was obtained. The appearance of types 1, 2, and 3 ice nuclei, catalyzing nucleation at -2 to -5, -5 to -7, and -7 to -10 degrees C, respectively, was investigated during low-temperature induction in wild-type strains and in a unique, detergent-sensitive mutant that contained no type 3 ice nuclei when grown at 32 degrees C. Nuclei appeared in the following order: type 3, then type 2 and type 1. The disappearance of classes of ice nuclei was monitored during high-temperature treatment of fully induced cells; nuclei disappeared in the order type 1, type 2, and type 3. Although analysis of nucleation events is complicated by masking and unmasking of ice sites in the same cells, these temporal sequences of ice nucleus appearance or disappearance are consistent with an aggregation model for ice nucleus assembly (A. G. Govindarajan and S. E. Lindow, Proc. Natl. Acad. Sci. USA 85:1334-1338, 1988; G. Warren and P. Wolber, Mol. Microbiol. 5:239-243, 1991).

Cell Membrane↗

PubMed1992

Isoprene emission rate and intercellular isoprene concentration as influenced by stomatal distribution and conductance.

Isoprene emission in relation to stomatal distribution and conductance was determined for the hypostomatous species, aspen and white oak, and the amphistomatous species, cottonwood. For aspen and oak, isoprene emission from the adaxial (nonstomatal) surface was <2% of that from the abaxial (stomatal) surface, even when stomata were closed by addition of abscisic acid (ABA). When treated with ABA, the total flux rate of isoprene emission from leaves of these two hypostomatous species was unchanged, despite decreases in stomatal conductance of over 90%. The lack of control over isoprene emission rate by stomatal conductance, despite the apparent movement of isoprene through the stomatal pores, was due to increases in the intercellular isoprene concentration that compensated for the decreased stomatal conductance and restored the equilibrium between the isoprene synthesis rate and emission rate. This relationship was demonstrated by (a) an experiment in which the decrease in the internal isoprene pool following the imposition of darkness took longer in the presence of ABA than in its absence, and (b) direct measurements of the internal isoprene concentration through vacuum extraction, which revealed substantially higher values in the presence of ABA than in its absence. In the amphistomatous species, cottonwood, isoprene was emitted from both surfaces and addition of ABA caused an increase in isoprene emission from one surface coupled with a decrease from the other surface. The specific surface exhibiting an increase varied among leaves, with some leaves exhibiting an increase from the adaxial surface and other leaves from the abaxial surface. We interpret this as indicating nonuniform stomatal closure with concomitant emission of isoprene at the greatest rate from the surface with the highest stomatal conductance. We also observed an increase in the total isoprene emission rate from cottonwood leaves following treatment with ABA. We interpret this as indicating a stimulation of isoprene synthesis in response to ABA or stomatal closure, with unknown cause.

Journal Article↗

PubMed1992

Relationships among Isoprene Emission Rate, Photosynthesis, and Isoprene Synthase Activity as Influenced by Temperature.

Isoprene emissions from the leaves of velvet bean (Mucuna pruriens L. var utilis) plants exhibited temperature response patterns that were dependent on the plant's growth temperature. Plants grown in a warm regimen (34/28 degrees C, day/night) exhibited a temperature optimum for emissions of 45 degrees C, whereas those grown in a cooler regimen (26/20 degrees C, day/night) exhibited an optimum of 40 degrees C. Several previous studies have provided evidence of a linkage between isoprene emissions and photosynthesis, and more recent studies have demonstrated that isoprene emissions are linked to the activity of isoprene synthase in plant leaves. To further explore this linkage within the context of the temperature dependence of isoprene emissions, we determined the relative temperature dependencies of photosynthetic electron transport, CO(2) assimilation, and isoprene synthase activity. When measured over a broad range of temperatures, the temperature dependence of isoprene emission rate was not closely correlated with either the electron transport rate or the CO(2) assimilation rate. The temperature optima for electron transport rate and CO(2) assimilation rate were 5 to 10 degrees C lower than that for the isoprene emission rate. The dependence of isoprene emissions on photon flux density was also affected by measurement temperature in a pattern independent of those exhibited for electron transport rate and CO(2) assimilation rate. Thus, despite no change in the electron transport rate or CO(2) assimilation rate at 26 and 34 degrees C, the isoprene emission rate changed markedly. The quantum yield of isoprene emissions was stimulated by a temperature increase from 26 to 34 degrees C, whereas the quantum yield for CO(2) assimilation was inhibited. In greenhouse-grown aspen leaves (Populus tremuloides Michaux.), the high temperature threshold for inhibition of isoprene emissions was closely correlated with the high temperature-induced decrease in the in vitro activity of isoprene synthase. When taken together, the results indicate that although there may be a linkage between isoprene emission rate and photosynthesis, the temperature dependence of isoprene emission is not determined solely by the rates of CO(2) assimilation or electron transport. Rather, we propose that regulation is accomplished primarily through the enzyme isoprene synthase.

Journal Article↗

PubMed1991

Delayed Onset of Isoprene Emission in Developing Velvet Bean (Mucuna sp.) Leaves.

Isoprene (2-methyl-1,3-butadiene) is one of the major volatile hydrocarbons emitted by plants, but its biosynthetic pathway and role in plant metabolism are unknown. Mucuna sp. (velvet bean) is an isoprene emitter, and leaf isoprene emission rate increased as much as 125-fold as leaves developed, and declined in older leaves. Net CO(2) assimilation and stomatal conductance, under different growth and environmental conditions, increased 3 to 5 days prior to an increase in isoprene emission rate, indicating that photosynthetic competence develops before significant isoprene emission occurs.

Journal Article↗

PubMed1991

Enzymatic synthesis of isoprene from dimethylallyl diphosphate in aspen leaf extracts.

Aspen (Populus tremuloides Michx.) leaf extracts contain a newly discovered enzyme activity that catalyzes the magnesium ion-dependent elimination of diphosphate from dimethylallyl diphosphate with rearrangement to form isoprene (2-methyl, 1-3-butadiene). This isoprene synthase activity has been partially purified. The nonenzymatic reaction of dimethylallyl diphosphate to isoprene, known to be acid catalyzed, may be insignificant at physiological pH. In contrast, the enzymatic reaction may be responsible for the majority of light-dependent isoprene production by isoprene-emitting plants.

Journal Article↗

PubMed1989

Isoprene emission from aspen leaves : influence of environment and relation to photosynthesis and photorespiration.

Isoprene emission rates from quaking aspen (Populus tremuloides Michx.) leaves were measured simultaneously with photosynthesis rate, stomatal conductance, and intercellular CO(2) partial pressure. Isoprene emission required the presence of CO(2) or O(2), but not both. The light response of isoprene emission rate paralleled that of photosynthesis. Isoprene emission was inhibited by decreasing ambient O(2) from 21% to 2%, only when there was oxygen insensitive photosynthesis. Mannose (10 millimolar) fed through cut stems resulted in strong inhibition of isoprene emission rate and is interpreted as evidence that isoprene biosynthesis requires either the export of triose phosphates from the chloroplast, or the continued synthesis of ATP. Light response experiments suggest that photosynthetically generated reductant or ATP is required for isoprene biosynthesis. Isoprene biosynthesis and emission are not directly linked to glycolate production through photorespiration, contrary to previous reports. Isoprene emission rate was inhibited by above-ambient CO(2) partial pressures (640 microbar outside and 425 microbar inside the leaf). The inhibition was not due to stomatal closure. This was established by varying ambient humidity at normal and elevated CO(2) partial pressures to measure isoprene emission rates over a range of stomatal conductances. Isoprene emission rates were inhibited at elevated CO(2) despite no change in stomatal conductance. Addition of abscisic acid to the transpiration stream dramatically inhibited stomatal conductance and photosynthesis rate, with a slight increase in isoprene emission rate. Thus, isoprene emission is independent of stomatal conductance, and may occur through the cuticle. Temperature had an influence on isoprene emission rate, with the Q(10) being 1.8 to 2.4 between 35 and 45 degrees C. At these high temperatures the amount of carbon lost through isoprene emission was between 2.5 and 8% of that assimilated through photosynthesis. This represents a significant carbon cost that should be taken into account in determining midsummer carbon budgets for plants that are isoprene emitters.

Journal Article↗

PubMed1988

Inhibition of bacterial ice nucleators by fish antifreeze glycoproteins.

Certain bacteria promote the formation of ice in super-cooled water by means of ice nucleators which contain a unique protein associated with the cell membrane. Ice nucleators in general are believed to act by mimicking the structure of an ice crystal surface, thus imposing an ice-like arrangement on the water molecules in contact with the nucleating surface and lowering the energy necessary for the initiation of ice formation. Quantitative investigation of the bacterial ice-nucleating process has recently been made possible by the discovery of certain bacteria that shed stable membrane vesicles with ice nucleating activity. The opposite effect, inhibition of ice formation, has been described for a group of glycoproteins found in different fish and insect species. This group of substances, termed antifreeze glycoproteins (AFGPs), promotes the supercooling of water with no appreciable effect on the equilibrium freezing point or melting temperature. Substantial evidence now indicates that AFGPs act by binding to a growing ice crystal and slowing crystal growth. As the ice-nucleating protein surface is believed to have a structure similar to an embryonic ice crystal, AFGPs might be predicted to interact directly with a bacterial ice-nucleating site. We report here that AFGPs from the antarctic fish Dissostichus mawsoni inhibit the ice-nucleating activity of membrane vesicles from the bacterium Erwinia herbicola. The inhibition effect shows saturation at high concentration of AFGP and conforms to a simple binding reaction between the AFGP and the nucleation centre.

Animals↗

PubMed1988

Purification and preliminary characterization of a monomeric glutathione S-transferase from Tetrahymena thermophila.

Cell extracts of the protozoan Tetrahymena thermophila contain high levels of glutathione S-transferase (EC 2.5.1.18). The level of the enzyme varied with the growth cycle, increasing in early stationary phase. It was localized in the cytoplasm. Only one major molecular form of the enzyme was detected by ion-exchange and gel filtration chromatography and isoelectric focusing (pI 7.0), and it was purified to homogeneity. The molecular weight of the enzyme was estimated to be 35,000 from gel filtration and 33,000 from sodium dodecyl sulfate-gel electrophoresis, indicating the enzyme is a monomer. The enzyme was specific for glutathione as the thiol substrate, and was most active with 1-chloro-2,4-dinitrobenzene as the electrophilic substrate. It also exhibited glutathione peroxidase activity. The apparent Km values for glutathione and 1-chloro-2,4-dinitrobenzene were 3.2 and 1.5 mM, respectively. The enzyme catalyzed an S-aryltransfer with pentachloronitrobenzene (Km = 7.2 microM) consistent with the view that this pesticide is metabolized in part via S-conjugation pathways in T. thermophila.

Animals↗

PubMed1987

High incidence of selenite-resistant bacteria from a site polluted with selenium.

The level of selenium-resistant bacteria in water, algal mats, and sediment from Kesterson reservoir, Calif., a site with known selenium pollution, was compared with that in nearby Volta reservoir, a site with low selenium levels. A high percentage (greater than 50%) of all isolates from the Kesterson samples were resistant to 10 mM selenite. In contrast, only a small percentage of the Volta isolates were resistant to this level of selenite. The identity of some selenite-resistant isolates and MICs of selenite, selenate, arsenate, tellurite, and tellurate were determined.

Bacteria↗

PubMed1987

Widespread occurrence of bacterial thiol methyltransferases and the biogenic emission of methylated sulfur gases.

A majority of heterotrophic bacteria isolated from soil, water, sediment, vegetation, and marine algae cultures methylated sulfide, producing methanethiol. This was demonstrated with intact cells by measuring the emission of methanethiol with a sulfur-selective chemiluminescence detector, and in cell extracts by detection of sulfide-dependent thiol methyltransferase activity. Extracts of two Pseudomonas isolates were fractionated by gel-filtration and ion-exchange chromatography, and with sulfide as the substrate a single peak of thiol methyltransferase activity was seen in each case. Extracts of several bacterial strains also contained thiol methyltransferase activity with organic thiols as substrates. Thus, S-adenosylmethionine-dependent thiol methyltransferase activities are widespread in bacteria and may contribute to biogenic emissions of methylated sulfur gases and to the production of methyl thioethers.

Bacteria, Aerobic↗

PubMed1987

Enzymatic methylation of sulfide, selenide, and organic thiols by Tetrahymena thermophila.

Cell extracts from the ciliate Tetrahymena thermophila catalyzed the S-adenosylmethionine-dependent methylation of sulfide. The product of the reaction, methanethiol, was detected by a radiometric assay and by a gas-chromatographic assay coupled to a sulfur-selective chemiluminescence detector. Extracts also catalyzed the methylation of selenide, and the product was shown by gas chromatography-mass spectrometry to be methaneselenol. The sulfide and selenide methyltransferase activities copurified with the aromatic thiol methyltransferase previously characterized from this organism (A.-M. Drotar and R. Fall, Pestic. Biochem. Physiol. 25:396-406, 1986), but heat inactivation experiments suggested the involvement of distinct sulfide and selenide methyltransferases. Short-term toxicity tests were carried out for sulfide, selenide, and their methylated derivatives; the monomethylated forms were somewhat more toxic than the nonmethylated or dimethylated compounds. Cell suspensions of T. thermophila exposed to sulfide, methanethiol, or their selenium analogs emitted methylated derivatives into the headspace. These results suggest that this freshwater protozoan is capable of the stepwise methylation of sulfide and selenide, leading to the release of volatile methylated sulfur or selenium gases.

Animals↗

PubMed1986

A differential scanning calorimeter for ice nucleation distribution studies--application to bacterial nucleators.

A differential scanning calorimeter has been developed for the automatic detection and measurement of dropwise freezing within a sample of 100-200 water drops. A typical drop size of 1 microliter is employed. The sample is distributed on flat, square (4-cm) thermoelectric sensors and the temperature is scanned downward by conductive cooling to a liquid nitrogen bath. The rate of cooling, typically 1 degree C/min, is set by the choice of a heat conduction rod between the calorimeter and the liquid nitrogen bath. The voltages from the thermopiles along with a system temperature-measuring thermocouple are continuously monitored by digital voltmeters and recorded every half-second in a computer memory. A freezing event in a drop is detected by a characteristic voltage signal whose integral with time is proportional to the size of the drop and its heat of fusion. The half-life of a freezing event signal is 10 s for a 1-microliter drop. The integrated signal produced from multiple freezing events is shown to provide a direct measure of the number of drops frozen at a given temperature. A distribution curve and its smoothed derivative can be constructed directly from these measurements. The instrument, which is termed an "ice nucleometer," is illustrated in determining the ice nucleation distribution in a population of Escherichia coli harboring cloned ice nucleation genes.

Calorimetry, Differential Scanning↗