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Biomedical subjects

F A Loewus

Publications and source records attributed to F A Loewus.

At least 37 records · Page 2Linked to original sources

Enantiomeric Form of myo-Inositol-1-Phosphate Produced by myo-Inositol-1-Phosphate Synthase and myo-Inositol Kinase in Higher Plants.

The product of myo-inositol-1-phosphate synthase, EC 5.5.1.4, from mature pollen of Lilium longiflorum Thunb., cv Ace (Easter lily) and that of myo-inositol kinase, EC 2.7.1.64, from wheat germ has been identified as 1l-myo-inositol-1-phosphate by gas chromatography of its trimethylsilyl-methyl phosphate derivative on a glass capillary column bearing a chiral phase.

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Sugar Uptake in Lily Pollen : A PROTON SYMPORT.

The data presented here are consistent with a proton-sugar co-transport in germinated pollen of Lilium longiflorum Thunb. Optimal uptake occurs at pH 5.0. A K(m) of 1.7 to 1.8 millimolar is obtained from the initial rate of pH change induced by sucrose uptake as well as from uptake of [U-(14)C]-sucrose. The energy of activation is - 11 kilocalories mole(-1). The effect of several inhibitors and sugar competitors on [U-(14)C]sucrose and d-[U-(14)C] glucose uptake is given. The possibility of hydrolysis of sucrose prior to its transport into the pollen tube has been considered and reasons for choosing a sucrose-type uptake are presented. The possible in vivo significance of this co-transport process during pollen germination is discussed. Germinated pollen has features to recommend it as an experimental system of choice for studies of sugar uptake.

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Stereochemistry of the myo-inositol-1-phosphate synthase reaction.

Experiments with D-glucose-6-P stereospecifically tritiated at C-6 showed that the myo-inositol-1 P synthase reaction catalyzed by both the enzyme from beef testis and from pollen of Lilium longiflorum proceeds with stereospecific loss of the pro-6R and incorporation of the pro-6S hydrogen into the product. The ring closure thus occurs in a retention mode at C-6 of the substrate, a finding at variance with an earlier report, but in agreement with the stereochemistry recently determined for the reaction in Streptomyces flavopersicus and with the general stereochemical mode of operation of aldolases.

Animals↗

The C-5 hydrogen isotope-effect in myo-inositol 1-phosphate synthase as evidence for the myo-inositol oxidation-pathway.

The hydrogen isotope-effect that occurs in vitro during myo-inositol1-phosphate synthase-catalyzed conversion of D-[5-3H]glucose 6-phosphate into myo[2-3H]inositol 1-phosphate has been used to compare the functional role of the nucleotide sugar oxidation-pathway with that of the myo-inositol oxidation-pathway in germinating lily pollen. Results reveal a significant difference between the 3H/14C ratios of glucosyl and galactosyluronic residues from pectinase-amyloglucosidase hydrolyzates of the 70% ethanol-insoluble fraction of D-[5-3H, 1-14C]glucose-labeled, germinating lily pollen. This isotope effect at C-5 of D-glucose that occurred during its conversion into D-galactosyluronic residues of pectic substance is not explained by loss of 3H when UDP-D-[5-3H, 1-14C]glucose is oxidized by UDP-D-glucose dehydrogenase from germinating lily pollen. The evidence obtained from this study favors a functional role for the myo-inositol oxidation-pathway during in vivo conversion of glucose into galactosyluronic residues of pectin in germinating lily pollen.

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Metabolism of myo-[2-H]Inositol and scyllo-[R-H]Inositol in Ripening Wheat Kernels.

Injection of myo-[2-(3)H]inositol or scyllo-[R-(3)H]inositol into the peduncular cavity of wheat stalks about 2 to 4 weeks postanthesis led to rapid translocation into the spike and accumulation of label in developing kernels, especially the bran fraction. With myo-[2-(3)H]inositol, about 50 to 60% of the label was incorporated into high molecular weight cell wall substance in the region of the injection. That portion translocated to the kernels was utilized primarily for cell wall polysaccharide formation and phytate biosynthesis. A small amount was recovered as free myo-inositol and galactinol. When scyllo-[R-(3)H]inositol was supplied, most of the label was translocated into the developing kernels where it accumulated as free scyllo-inositol and O-alpha-d-galactopyranosyl-scyllo-inositol in approximately equal amount. None of the label from scyllo-[R-(3)H]inositol was utilized for either phytate biosynthesis or cell wall polysaccharide formation.

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Metabolic Studies on Intermediates in the myo-Inositol Oxidation Pathway in Lilium longiflorum Pollen: III. Polysaccharidic Origin of Labeled Glucose.

On the basis of solubility, hydrolysis by glucoamylase (EC 3.2.1.3), and monomeric composition, starch appears to be the major glucose-containing, hot water-soluble polysaccharide that is labeled when germinated lily (Lilium longiflorum Thunb., cv. Ace) pollen is grown in the presence of myo-[2-(3)H]inositol, d-[R5,S5-(3)H]xylose, or l-[1-(14)C]arabinose.

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Metabolic Studies on Intermediates in the myo-Inositol Oxidation Pathway in Lilium longiflorum Pollen: I. Conversion to Hexoses.

The myo-inositol oxidation pathway was investigated in regard to its role as a source of carbon for products of hexose monophosphate metabolism in germinated pollen of Lilium longiflorum Thunb., cv. Ace. myo-[2-(14)]Inositol and d-[1-(14)C]glucuronate had similar distributions of radioactivity, contributing about three times more label to polysaccharide-bound glucose than myo-[2-(3)H]inositol. In the course of glucogenesis label from the latter appeared as tritiated water in the medium. This exchange could be enhanced by supplying d-[5R,5S-(3)H]xylose instead of myo-[2-(3)H]inositol. When the former was administered, [(3)H]glucose was the only labeled sugar residue found in polysaccharide products. The soluble constituents of d-[5R,5S-(3)H]xylose-labeled pollen contained no traces of labeled xylose despite massive uptake and utilization.l-[1-(14)C]- and l-[5-(14)C]Arabinose produced similar labeling patterns in germinated pollen including incorporation of arabinosyl units into pollen tube polysaccharides and substantial glucogenesis which led to utilization of arabinose for respiration and further incorporation of labeled glucosyl units into pollen tube polysaccharides.d-[5-(3)H]Galacturonate was rapidly taken up by germinated pollen but slowly utilized, without conversion to other sugars, for incorporation into pollen tube polysaccharides. l-[6-(14)C]Gulonate was not taken up by pollen.Results strongly support a scheme of conversion from myo-inositol to hexose monophosphate and subsequent products of glucose metabolism that involves the myo-inositol oxidation pathway.

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Metabolic Studies on Intermediates in the myo-Inositol Oxidation Pathway in Lilium longiflorum Pollen: II. Evidence for the Participation of Uridine Diphosphoxylose and Free Xylose as Intermediates.

myo-Inositol-linked glucogenesis in germinated lily (Lilium longiflorum Thunb., cv. Ace) pollen was investigated by studying the effects of added l-arabinose or d-xylose on metabolism of myo-[2-(3)H]inositol and by determining the distribution of radioisotope in pentosyl and hexosyl residues of polysaccharides from pollen labeled with myo-[2-(14)C]inositol, myo-[2-(3)H]inositol, l-[5-(14)C]arabinose, and d-[5R,5S-(3)H]xylose.myo-[2-(14)C]Inositol and l-[5-(14)C]arabinose produced labeled glucose with similar patterns of distribution of (14)C, 35% in C1, and 55% in C6. Arabinosyl units were labeled exclusively in C5. Incorporation of (3)H into arabinosyl and xylosyl units in pollen labeled with myo-[2-(3)H]inositol was repressed when unlabeled l-arabinose was included in the germination medium and a related (3)H exchange with water was stimulated. Results are consistent with a process of glucogenesis in which the myo-inositol oxidation pathway furnishes UDP-d-xylose as a key intermediate for conversion to hexose via free d-xylose and the pentose phosphate pathway.Additional evidence for this process was obtained from pollen labeled with d-[5R,5S-(3)H]xylose or myo-[2-(3)H]inositol which produces d-[5R-(3)H]xylose. Glucosyl units from polysaccharides in the former had 11% of the (3)H in C1 and 78% in C6 while glucosyl units in the latter had only 4% in C1 and 78% in C6. Stereochemical considerations involving selective exchange with water of prochiral-R (3)H in C1 of fructose-6-P during conversion to glucose provide explanation for observed differences in the metabolism of these 5-labeled xyloses.Incorporation of (3)H from myo-[2-(3)H]inositol into arabinosyl and xylosyl units of pollen polysaccharides was unaffected by the presence of unlabeled d-xylose in the medium. Exchange of (3)H with water was greatly affected, decreasing from a value of 21% exchange in the absence of unlabeled d-xylose to 5% in the presence of 6.7 mmd-xylose.d-Xylose was rapidly utilized for glucogenesis by germinated pollen tubes. This observation supports the view that free d-xylose is an important intermediate following breakdown of UDP-d-xylose during myo-inositol-linked glucogenesis.

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Further Studies on Oxalic Acid Biosynthesis in Oxalate-accumulating Plants.

l-Ascorbic acid functions as a precursor of oxalic acid in several oxalate-accumulating plants. The present study extends this observation to include Rumex crispus L. (curly dock), Amaranthus retroflexus L. (red root pigweed), Chenopodium album L. (lamb's-quarters), Beta vulgaris L. (sugar beet), Halogeton glomeratus M. Bieb. (halogeton), and Rheum rhabarbarum L. (rhubarb). Several species with low oxalate content are also examined.When l-[1-(14)C]ascorbic acid is supplied to young seedlings of R. crispus or H. glomeratus, a major portion of the (14)C is released over a 24-hour period as (14)CO(2) and only a small portion is recovered as [(14)C]oxalate, unlike cuttings from 2- or 4-month-old plants which retain a large part of the (14)C as [(14)C]oxalic acid and release very little (14)CO(2). Support for an intermediate role of oxalate in the release of (14)CO(2) from l-[1-(14)C]ascorbic acid is seen in the rapid release of (14)CO(2) by R. crispus and H. glomeratus seedlings labeled with [(14)C]oxalic acid.The common origin of oxalic acid carbon in the C1 and C2 fragment from l-ascorbic acid is demonstrated by comparison of (14)C content of oxalic acid in several oxalate-accumulators after cuttings or seedlings are supplied equal amounts of l-[1-(14)C]- or l-[UL-(14)C]ascorbic acid. Theoretically, l-[1-(14)C]ascorbic acid will produce labeled oxalic acid containing three times as much (14)C as l-[UL-(14)C]ascorbic acid when equal amounts of label are provided. Experimentally, a ratio of 2.7 +/- 0.5 is obtained in duplicate experiments with six different species.

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Biosynthesis of (+)-Tartaric Acid from l-[4-C]Ascorbic Acid in Grape and Geranium.

The metabolic fate of l-[4-(14)C]ascorbic acid has been examined in the grape (Vitis labrusca L.) and lemon geranium (Pelargonium crispum L. L'Hér. cv. Prince Rupert) under conditions comparable to data from l-[1-(14)C]ascorbic acid and l-[6-(14)C]ascorbic acid experiments. In detached grape leaves and immature berries, l-[4-(14)C]ascorbic acid and l-[1-(14)C]ascorbic acid were equivalent precursors to carboxyl labeled (+)-tartaric acid. In geranium apices, l-[4-(14)C]ascorbic acid yielded internal labeled (+)-tartaric acid while l-[6-(14)C]ascorbic acid gave an equivalent conversion to carboxyl labeled (+)-tartaric acid. These findings clearly show that two distinct processes for the synthesis of (+)-tartaric acid from l-ascorbic acid exist in plants identified as (+)-tartaric acid accumulators. In grape leaves and immature berries, (+)-tartaric acid synthesis proceeds via preservation of a four-carbon fragment derived from carbons 1 through 4 of l-ascorbic acid while carbons 3 through 6 yield (+)-tartaric acid in geranium apices.

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Evidence for a Functional myo-Inositol Oxidation Pathway in Lilium longiflorum Pollen.

Addition of myo-inositol to pentaerythritol-based germination media repressed the conversion of d-[1-(14)C]glucose to labeled uronosyl and pentosyl units of tube wall pectic substance in lily pollen (Lilium longiflorum Thunb.). Conversion of d-[1-(14)C]glucose to labeled glucosyl, galactosyl, and rhamnosyl units was unaffected. The reverse experiment, addition of d-glucose to pentaerythritol-based media, failed to affect the conversion of myo-[2-(3)H]inositol to uronosyl and pentosyl units although the flow of label into products of myo-inositol-linked glucogenesis was blocked. Results of these experiments are discussed in terms of a functional myo-inositol oxidation pathway.d-[1-(14)C]Glucose-labeled pollen tubes contain a labeled, 70% ethyl alcohol-soluble, acidic compound whose formation is blocked by the myo-inositol antagonist, 2-O,C-methylene-myo-inositol (Chen et al. 1977 Plant Physiol. 59: 658). This compound has been identified as l-malic acid.

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myo-Inositol Metabolism in Lilium longiflorum Pollen: Uptake and Incorporation of myo-Inositol-2-H.

Germinating Lilium longiflorum pollen absorbs and metabolizes myo-inositol-2-(3)H (MI-2-(3)H) with a pronounced lag when label is supplied from the beginning of germination. If MI-2-(3)H is given after 3 hours of germination, incorporation of labeled metabolic products into pollen tube polysaccharides is constant over a range of 0.56 mm to 2.78 mm MI. When MI-2-(3)H is supplied as a 0.5-hour pulse 3 hours after germination, the proper precursor-product relationship to tube wall polysaccharides is observed. Replacing 10% of the germination media with sigmatic exudate from a compatible cultivar hastens germination and tube elongation. Enhanced MI metabolism accompanies tube growth in this exudate-enriched media.

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Effect of a myo-Inositol Antagonist, 2-O, C-Methylene-myo-Inositol, on the Metabolism of myo-Inositol-2-H and d-Glucose-1-C in Lilium longiflorum Pollen.

2-O,C-Methylene-myo-inositol (MMO), a myo-inositol (MI) antagonist, inhibits germination and tube elongation of pollen from Lilium longiflorum cv. Ace or 44. The presence of 5 mm MMO in Dickinson's pentaerythritol medium (Plant Physiol. 43:1-8) partially blocks germination. The tubes produced are short and fail to elongate. In the presence of MI, MMO's toxic effect is blocked. As little as 0.56 mm MI will maintain normal germination in the presence of 43 mm MMO, and pollen tubes continue to elongate for 2 to 3 hr. Eventually, the toxic action of MMO prevents further growth. MMO does not inhibit UDP-d-glucose dehydrogenase from lily pollen.Uptake of MI-2-(3)H and incorporation of tritium into galacturonic acid and pentose units of tube wall pectin are blocked by MMO. The site of this inhibition is undertermined. Uptake of d-glucose-1-(14)C and incorporation of (14)C into 70% ethyl alcohol-insoluble polysaccharides of germinated pollen are not blocked by MMO, but distribution of label into polysaccharide product is altered. In MMO-treated pollen, very little (14)C is found in uronic acid or pentose units. At 30 mm MMO, about two-thirds of the carbon flow from d-glucose to these pectic components is interrupted. MMO also alters d-glucose metabolism in the 70% ethyl alcohol-soluble fraction, but the compound involved must still be identified.These results offer fresh evidence of an intermediary role for MI during UDP-d-glucuronate biosynthesis in germinated pollen.

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Effect of Benzyladenine, 2,4-Dichlorophenoxyacetic Acid, and d-Glucose on myo-Inositol Metabolism in Acer pseudoplatanus L. Cells Grown in Suspension Culture.

Suspension cultures of Acer pseudoplatanus L. cells grown for 15 days in medium (T. Murashige and F. Skoog. 1962. Physiol. Plant. 15: 473-497) contained 3% sucrose, 1 mg/l 6-benzylaminopurine (BA), and 0.1 mg/l 2,4-dichlorophenoxyacetic acid (2,4-D), referred to here as normal media, removed newly added myo-inositol-2-(3)H up to 100 mg/l in 24 hours and utilized up to 20% of this cyclitol for pectin biosynthesis. When the BA content of the growth medium was raised 10-fold, uptake of myo-inositol was drastically reduced and very little was available for pectin biosynthesis. Neither cell growth as measured by packed cell volume or by dry weight, nor monomer composition of pectic polysaccharides was affected by the increased level of cytokinin. Increasing, 2,4-d 10-fold instead of BA had little or no effect on myo-inositol uptake, although it did reduce the amount of myo-inositol utilized for pectin biosynthesis. Cells grown 15 days in normal media failed to remove added myo-inositol if 3% d-glucose was included. The net result was similar to that found in cells grown in the high BA condition. If a trace amount of d-galactose-1-(14)C was supplied to cells after 15 days of growth in normal, high BA, or high 2,4-d media, there was no significant variation in uptake and utilization of label among the three growth conditions.

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Stereoisomeric Characterization of Tartaric Acid Produced during l-Ascorbic Acid Metabolism in Plants.

Labeled tartaric acids from Pelargonium crispum apices which had been fed l-ascorbic acid-6-(14)C and Vitis labrusca and Parthenocissus inserta tissues which had been fed l-ascorbic acid-1-(14)C were examined by chemical means to determine chiral configuration. In each instance, label was associated with (+)-tartaric acid.Similar experiments with labeled tartaric acid from P. crispum which had been labeled with d-glucose-1-(14)C or -6-(14)C led to the same result. No evidence was obtained for formation of labeled meso-tartaric acid in experiments described above. The recent suggestion of H. Ruffner and D. Rast (Z. Pflanzenphysiol. 73: 45-55, 1974) that conversion of l-ascorbic acid to tartaric acid in plants is a nonenzymatic process is re-examined in the light of present findings.

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Metabolic Conversion of l-Ascorbic Acid to Oxalic Acid in Oxalate-accumulating Plants.

l-Ascorbic acid-1-(14)C and its oxidation product, dehydro-l-ascorbic acid, produced labeled oxalic acid in oxalate-accumulating plants such as spinach seedlings (Spinacia oleracea) and the detached leaves of woodsorrel (Oxalis stricta and O. oregana), shamrock (Oxalis adenopylla), and begonia (Begonia evansiana). In O. oregana, conversion occurred equally well in the presence or absence of light. This relationship between l-ascorbic acid metabolism and oxalic acid formation must be given careful consideration in attempts to explain oxalic accumulation in plants.

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