Polyamine distributions in thermophilic eubacteria belonging to Thermus and Acidothermus

Triamines such as norspermidine, spermidine, and homospermidine and tetraamines such as norspermine, spermine, thermospermine, and aminopropylhomospermidine were found to be distributed ubiquitously in the eight extremely thermophilic (growing at 70 degrees C) Thermus species tested. Three linear pentaamine (caldopentamine, homocaldopentamine, and thermopentamine), two linear hexaamines (caldohexamine and homocaldohexamine), two tertiary branched tetraamines (N4-aminopropylnorspermidine and N4-aminopropyl-spermidine), and quaternary branched pentaamines such as N4-bis(aminopropyl)norspermidine and N4-bis(aminopropyl)spermidine were detected in T. thermophilus HB8, T. filiformis Wai33 A1, T. flavus AT-62, and T. caldophilus GK24. The linear hexaamines and branched polyamines were absent in T. aquaticus YT-1, T. sp. X-1, T. sp. T2, and T. sp. T351, in which linear pentaamines were minor components. Moderately thermophilic Thermus ruber and Thermus sp. K-2 contained putrescine, spermidine, norspermidine, homospermidine, spermine, norspermine, thermospermine, and aminopropylhomospermidine. No pentaamines, hexaamines, or branched polyamines were found in these two moderately thermophilic Thermus species. On the other hand, moderately thermophilic, acidophilic Acidothermus cellulolyticus was devoid of all the polyamines.     

Polyamine distribution profiles in the eighteen genera phylogenetically located within the Flavobacterium-Flexibacter-Cytophaga complex

Cellular polyamines of eighteen genera belonging to the Flavobacterium-Flexibacter-Cytophaga complex were analysed by ion exchange liquid chromatography. Homospermidine was the major polyamine in the genera Bergeyella, Riemerella, Ornithobacterium, Weeksella, Capnocytophaga, Polaribacter and Psychroflexus belonging to the family Flavobacteriaceae. In the family Spirosomaceae, Runella, Spirosoma and Flectobacillus species contained spermidine whereas Cyclobacterium species contained homospermidine. Within a divergent cluster, Haliscomenobacter and Lewinella species contained spermidine whereas Saprospira grandis contained agmatine alone. The major polyamine of Chitinophaga and Sporocytophaga species was homospermidine. Flexithrix dorotheae contained spermidine. Microscilla marina, the type species of the genus Microscilla, contained spermidine and cadaverine. However, 'Microscilla sericea' contained homospermidine, 'Microscilla furvescens' contained spermidine, and 'Microscilla arenaria' lacked all polyamines. Polyamine profiles serve as a phenotypic chemotaxonomic marker for the reclassification of the genera belonging to the complex.     

Polyamine Metabolism in the Roots of Phaseolus vulgaris. Interaction of the Inhibitors of Polyamine Biosynthesis with Putrescine in Growth and Polyamine Biosynthesis

The effects of the inhibitors of polyamine biosynthesis, canavanineand -methyl ornithine on growth, the activities of argininedecarboxylase (EC 4.1.1.19 [EC] ) and ornithine decarboxylase (EC4.1.1.17 [EC] ) and on polyamine content were examined in two differentgrowth regions of Phaseolus vulgaris L. cv. Taylor's Horticulturalroots. Separately, in the same manner, in the same bean rootsystem exogenous putrescine effect and the interaction of canavaninewith putrescine were determined. The arginine and ornithine decarboxylase activities found inroot apex were high where cell division activity was highest.Polyamine (putrescine and spermine) content did not correlatewith these activities, but polyamine level was high in the rootbase where cell elongation is the main process. The arginineanalogue, canavanine, inhibited arginine decayboxylase activityand polymine liters. Putrescine partially reversed the canavanineinhibition of root growth as well as arginine decarboxylaseactivity and polyamine content. Similarly -methyl ornithineslightly inhibited the root length and ornithine decarboxylaseactivity in the root apex. Besides, exogenous putrescine didnot effect significantly the endogenous polyamine titers. Theseresults reinforce the growing connection between polyaminesand the rates of cell devision in the roots of bean plants.Separately, arginine decarboxylase is the main enzyme in thebean roots. (Received November 10, 1986; Accepted March 3, 1987)     

Polyamine distribution profiles in newly validated genera and species within the Flavobacterium-Flexibacter-Cytophaga-Sphingobacterium complex

Cellular polyamines of 58 strains belonging to the Flavobacterium-Flexibacter-Cytophaga-Sphingobacterium complex were analysed by HPLC. Homospermidine was found in all species of Flavobacterium, Chryseobacterium, Empedobacter, Myroides, Cellulophaga, Salegentibacter, Psychroserpens and Gelidibacter of the family Flavobacteriaceae. Flavobacterium ferrugineum located outside of this family also contained homospermidine. Cytophaga fermentans and C. xylanolytica belonging to the family Bacteroidaceae contained spermidine. Cytophaga marinoflava and C. latercula belonging to Flavobacteriaceae contained homospermidine. The Cytophaga hutchinsonii/C. aurantiaca group contained homospermidine which was the major polyamine in Flexibacter maritimus/ F. ovolyticus of the family Flavobacteriaceae. The Flexibacter sancti/F filiformis/ Cytophaga arvensicola group, F. elegans, F. ruber, F. canadensis, F. flexilis and F. tractuosus, were located separately in different six clusters, and contained homospermidine. The Flexibacter litoralis/F. polymorphus/F. aggregans group contained spermidine, which was detected in Flexibacter roseolus belonging to a divergent cluster. Sphingobacterium and Pedobacter species of the family Sphingobacteriaceae contained homospermidine. Polyamine profiles serve, as a phenotypic chemotaxonomic marker, for the classification of this complex.     

Evolution of complex probability distributions in enzyme cascades

Unusual probability distribution profiles, including transient multi-peak distributions, have been observed in computer simulations of cell signaling dynamics. The emergence of these complex distributions cannot be explained using either deterministic chemical kinetics or simple Gaussian noise approximation. To develop physical insights into the origin of complex distributions in stochastic cell signaling, we compared our approximate analytical solutions of signaling dynamics with the exact numerical simulations. Our results are based on studying signaling in 2-step and 3-step enzyme amplification cascades that are among the most common building blocks of cellular protein signaling networks. We have found that while the multi-peak distributions are typically transient, and eventually evolve into single peak distributions, in certain cases these distributions may be stable in the limit of long times. We also have shown that introducing positive feedback loops results in diminution of the probability distribution complexity.     

Polyamine metabolism in the Microsporidia

Members of the phylum Microspora are all obligate intracellular parasites. Little is known concerning metabolic pathways in these parasites, some of which pose serious problems in immunocompromised patients. We investigated polyamine metabolism in the systemic pathogen Enterocytozoon cuniculi using intact pre-emergent spores, and cell-free preparations. We found both polyamine synthetic and interconversion pathways to be operative, as evidenced by conversion of ornithine into polyamines, and production of spermidine from spermine by pre-emergent spores. Recent developments in the antitumour field have highlighted the ability of bis-ethylated polyamine analogues to reduce polyamine levels and block growth of tumour cells. In light of enhanced polyamine uptake in Enc. cuniculi, we have begun to study bis-aryl 3-7-3 and bis-ethyl oligoamine analogues as leads for chemotherapy of microsporidia.     

Polyamine auxotrophs of Saccharomyces cerevisiae.

Strains of yeast have been constructed that are unable to synthesize ornithine and are thereby deficient in polyamine biosynthesis. These strains were used to develop a protocol for isolation of mutants blocked directly in polyamine synthesis. There were seven mutants isolated that lack ornithine decarboxylase activity; these strains exhibited greatly decreased pool levels of putrescine, spermidine, and spermine when grown in the absence of polyamines. Three of the mutants lack S-adenosylmethionine decarboxylase activity; polyamine limitation of a representative mutant resulted in an accumulation of putrescine and a decrease in spermidine and spermine. When the mutants were cultured in the absence of polyamines, a continuously declining growth rate was observed.     

Polyamine induced aggregation of DNA.

Polyamine induced aggregation of various DNAs has been studied under conditions usually employed in many enzymatic assays where DNA is one of the substrates. Spermine was by far the most efficient polyamine in causing aggregation followed by spermidine and cadaverine. All double-stranded and naturally occurring single-stranded DNAs were found to aggregate. No aggregation of single-stranded homodeoxypolymers could be detected under the same conditions. The concentration of polyamine at which the aggregation commenced was found to be a linear function of the DNA concentration. The slope of the curves depended on the nature of the polyamine, DNA the concentration of Mg++ and the ionic strength.     

Polyamine degradation in foetal and adult bovine serum.

1. Using protein-separative chromatographic procedures and assays specific for putrescine oxidase and spermidine oxidase, adult bovine serum was found to contain a single polyamine-degrading enzyme with substrate preferences for spermidine and spermine. Apparent Km values for these substrates were approx. 40 microM. The apparent Km for putrescine was 2 mM. With spermidine as substrate, the Ki values for aminoguanidine (AM) and methylglyoxal bis(guanylhydrazone) (MGBG) were 70 microM and 20 microM respectively. 2. Bovine serum spermidine oxidase degraded spermine to spermidine to putrescine and N8-acetylspermidine to N-acetylputrescine. Acrolein was produced in all these reactions and recovered in quantities equivalent to H2O2 recovery. 3. Spermidine oxidase activity was present in foetal bovine serum, but increased markedly after birth to levels in adult serum that were almost 100 times the activity in foetal bovine serum. 4. Putrescine oxidase, shown to be a separate enzyme from bovine serum spermidine oxidase, was present in foetal bovine serum but absent from bovine serum after birth. This enzyme displayed an apparent Km for putrescine of 2.6 microM. The enzyme was inhibited by AM and MGBG with Ki values of 20 nM. Putrescine, cadaverine and 1,3-diaminopropane proved excellent substrates for the enzyme compared with spermidine and spermine, and N-acetylputrescine was a superior substrate to N1- or N8-acetylspermidine.     

Polyamine Titre in Relation to Chill-Sensitivity in Phaseolus sp.

Guye, M G., Vigh, L. and Wilson, J. M. 1986. Polyamine titrein relation to chill-sensitivity in Phaseolus sp.—J. exp.Bot. 37: 1036–1043. Endogenous levels of the polyamines putrescine, spermidine andspermine were quantified in the primary leaves of five cultivarsof bean (Phaseolus sp.) differing in their ‘wilting response’to a chilling exposure of 5 ?C for 24 h. Levels of polyamines prior to chilling treatment did not appearto be correlated with chill-tolerance as levels in the non-chilledcontrols were highest in cultivars of medium chill-sensitivity.Plants grown under a vapour saturation deficit (VSD) of 8?4gm^–3 day/6?1 g m^–3 night exhibited a mild hardeningas compared to plants grown under a VSD of 5?7 gm^–3 day/4?1gm^–3 night, as the former showed less wilting on chilling.Hardening at high VSD had the effect of slightly lowering theputrescine content of non-chilled tissue but total polyaminecontent remained unchanged. However, on chilling, the largestrelative increase in polyamine levels, in particular that ofputrescine, occurred in hardened plants. There was also a significantrelative increase in putrescine titre in response to chillingin non-hardened genotypes of high chill-tolerance, whereas morechill-sensitive genotypes remained unchanged or slightly declinedin putrescine content on chilling. Relative changes in putrescine content rather than absolutelevels appears to be correlated with chill-tolerance. Theseresults are discussed in view of present knowledge on the adaptivesignificance of stress-induced changes in polyamines, especiallywith regard to membrane stability Key words: Chilling, polyamines, Phaseolus sp.     

Polyamine metabolism in enterocytes isolated from newborn pigs.

In the pig, the growth of intestinal mucosa is very intense after birth. Since the polyamines are key elements affecting cell proliferation and differentiation, the present work was undertaken in order to know whether this hypertrophy is associated with an adaptation of polyamine metabolism. Villus enterocytes isolated from pig immediately after birth or 2 days later were found to contain similar amounts of putrescine, spermidine and spermine, i.e., 0.23; 0.41 and 1.24 nmol/10(6) cells, respectively. At birth, despite a relatively high ODC activity, putrescine synthesis from 1 mM L-arginine or 2 mM L-glutamine was very low in isolated enterocytes (6.4 +/- 3.8 pmol/10(6) cells per 30 min), while spermidine and spermine production were not detectable. This could be explained by a very low L-ornithine generation from both amino acids and to an inhibitory effect of polyamines on ODC activity. Two days later, polyamine synthesis from L-arginine remained undetectable despite a higher L-ornithine generation. This was concomitant with a dramatic fall in ODC activity. At both stages, enterocytes were able to take up polyamines from the extracellular medium in a temperature-dependent manner. It is concluded that de-novo synthesis of polyamines from L-arginine or L-glutamine does not play a significant role in the control of polyamine content of pig enterocytes during the postnatal period. In contrast, polyamine uptake by enterocytes would contribute to maintain a steady-state polyamine content during this period.     

Polyamine toxicity in Neurospora crassa: protective role of the vacuole.

We used mutant strains of Neurospora crassa to define the discretionary capacity of this species for excess putrescine. The spe-3 mutant, which accumulates putrescine internally, and the puu-1 mutant, which accumulates toxic levels of putrescine from the medium, both sequestered large excesses of putrescine in vacuoles. Concomitantly in puu-1, inorganic polyphosphate increased modestly and some of the monovalent cation of the vacuole was discharged. These two factors contribute to the increased capacity for polyamines in this fungus. Putrescine, however, can exceed the capacity of vacuoles such that they no longer protect the cytosol from toxic levels of the amine. The puu-1 mutant illustrates the importance of the sequestration of intracellular polyamines, as well as the control of polyamine uptake through the cell membrane.     

Polyamine biosynthesis in the developing rabbit palate

Levels of putrescine, spermidine, and spermine and their biosynthetic enzymes, ornithine decarboxylase (ODC) and S-adenosylmethionine decarboxylase (SAMDC) were measured in the developing rabbit palate between day 14 and day 18 of gestation. DNA, RNA, and protein synthesis were also measured during this time period to determine if a correlation exists between polyamine biogenesis and macromolecular synthesis. ODC activity was found to be twice as high on day 14 as on the succeeding days of gestation, while SAMDC activity did not change significantly. Levels of putrescine and spermine were higher on day 14 by 22% and 30%, respectively, than levels on day 18. Spermidine concentration did not change. DNA synthesis remained relatively constant between days 14 and 18 of gestation, suggesting that there is no peak in cell proliferation during this period. RNA synthesis was elevated significantly on day 14 and protein synthesis was significantly higher on both days 14 and 16. This data indicates that there is no correlation between polyamine synthesis and cell proliferation during this period of palatal development, but polyamines could play a regulatory role in RNA and/or protein synthesis.     

Polyamine biosynthesis during germination of yeast ascospores.

The role of the diamine putrescine during germination and outgrowth of ascospores of Saccharomyces cerevisiae was examined. Ornithine decarboxylase activity increased and declined rapidly during germination and outgrowth; peak activity was attained after the cells had proceeded through the G1 interval of the cell cycle, whereas minimal activity was present at the completion of the first cell division. alpha-Methylornithine inhibited both ornithine decarboxylase activity and the in vivo accumulation of putrescine. In the presence of alpha-methylornithireak dormancy and proceed through one cell division. Subsequent cellular growth, however, was retarded but not completely inhibited. The supplementation of Methylglyoxal bis(guanylhydrazone) to sporulation medium greatly inhibited this sexual process. These data suggest that the synthesis of putrescine is not required for the breaking of spore dormancy, but that polyamine biosynthesis may be essential for meiosis and sporulation.     

植物源食品多胺的作用及其含量调节

多胺 (polyamines,PAs)在植物源食品中广泛分布 ,在人体健康中发挥着重要作用。简要概述了多胺在细胞增殖、肿瘤发展中的作用及其作用的初步机理。通过化学调控和基因工程手段以及一些环境条件的控制 ,可以人为地调节植物源食品多胺的含量。     

Polyamine biosynthesis during somatic embryogenesis in interior spruce (Picea glauca x Picea engelmannii complex)

Putrescine, spermidine, and spermine levels during somatic embryogenesis of interior spruce (Picea glauca x Picea engelmannii complex) were quantified On abscisic acid supplemented growth medium putrescine and spermidine levels increased two-fold coinciding with maturation of the early somatic embryos to globular embryos. Polyclonal antibodies raised against Escherichia coli arginine decarboxylase (ADC) and ornithine decarboxylase (ODC), following affinity purification specifically recognized spruce ADC and ODC, which corresponded to 85kD and 65kD bands on western blots of total protein extracts from embryogenic masses, Immunoassays using these antibodies showed increased ADC levels corresponding to embryo maturation while ODC levels remained the same. From these results it is concluded that polyamines are involved in the maturation of somatic embryos of interior spruce.Abbreviations ADC arginine decarboxylase - BSA bovine serum albumin - ODC ornithine decarboxylase - PBS phosphate buffered saline - PCA perchloric acid - SDS-PAGE sodium dodecyl sulfateporyacrylamide gel electrophoresis