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1.
Developmental change in c(6)-aldehyde formation by soybean leaves   总被引:3,自引:1,他引:2       下载免费PDF全文
Damage to plant leaves by wounding or freezing induces the production of large amounts of C6-compounds. However, the control of formation of these compounds in leaves is not yet clear. In the current study, C6-aldehyde formation by freeze-injured soybean leaves of different ages (based on the leaf positions on the plant) at stage R1 of plant development was investigated. The results demonstrate that C6-aldehyde formation by the soybean (Glycine max L.) leaves changes as leaves develop. Younger leaves produce high levels of C6-aldehydes, mainly composed of hexanal. Subsequently, as the leaves develop, the level of C6-aldehyde formation decreases markedly, followed by an increase with a large shift from hexanal to hexenals. Lipoxygenase and lipolytic acyl hydrolase activity was reduced, and, in contrast, hydroperoxide lyase activity increased. There was little difference in lipoxygenase substrate specificity for linoleic acid and linolenic acid, but hydroperoxide lyase preferentially utilized 13-hydroperoxy-9,11,15-octadecatrienoic acid. In the in vivo lipoxygenase substrate pool, the linoleic acid level declined and the relative level of linolenic acid increased. The change in ratios of linolenic acid to linoleic acid showed a similar trend during soybean leaf development to that of hexenals to hexanal.  相似文献   

2.
The homogenate of tea seed cotyledons contained an inhibitor for C6-aldehyde formation from linoleic acid and linolenic acid by isolated tea chloroplasts. Seed homogenates of other plants, such as soybean, kidney bean, cucumber, Japanese radish and rice, also contained the inhibitor for C6-aldehyde formation. The inhibitor from tea seed and cucumber seed inhibited C6-aldehyde formation by the homogenate of cucumber hypocotyl. Hydroperoxides of linoleic acid detected were reduced when the tea seed inhibitor was added to the reaction mixture, but the enzyme activities of lipoxygenase and hydroperoxide lyase were not inhibited. This means that the inhibitor is a decomposer of fatty acid hydroperoxides as an intermediate of C6-aldehyde formation. The tea seed inhibitor was formed during the seed ripening and it was stable during the seed germination. These findings obtained here suggest that the inhibitor is widely present in plant seeds and inhibits C6-aldehyde formation by a variety of plant tissues.  相似文献   

3.
1. Etiolated seedlings of alfalfa and cucumber evolved n-hexanal from linoleic acid and cis-3-hexenal and trans-2-hexenal from linolenic acid when they were homogenized.

2. The activities for n-hexanal formation from linoleic acid, lipoxygenase and hydro-peroxide lyase were maximum in dry seeds and 1~2 day-old etiolated seedlings of alfalfa, and in 6~7 day-old etiolated seedlings of cucumber.

3. n-Hexanal was produced from linoleic acid and 13-hydroperoxylinoleic acid by the crude extracts of etiolated alfalfa and cucumber seedlings. cis-3-Hexenal and trans-2-hexenal were produced from linolenic acid and 13-hydroperoxylinolenic acid by the crude extracts of etiolated alfalfa and cucumber seedlings. But these extracts, particulariy cucumber one, showed a high isomerizing activity from cis-3-hexenal to trans-2-hexenal.

4. When the C8-aldehydes were produced from linoleic acid and linolenic acid by the crude extracts, formation of hydroperoxides of these C18-fatty acids was observed.

5. When 9-hydroperoxylinoleic acid was used as a substrate, trans-2-nonenal was produced by the cucumber homogenate but not by the alfalfa homogenate.

6. As the enzymes concerned with C6-aldehyde formation, lipoxygenase was partially purified from alfalfa and cucumber seedlings and hydroperoxide lyase, from cucumber seedlings. Lipoxygenase was found in a soluble fraction, but hydroperoxide lyase was in a membrane bound form. Alfalfa lipoxygenase catalyzed formation of 9- and 13-hydroperoxylinoleic acid (35: 65) from linoleic acid and cucumber one, mainly 13-hydroperoxylinoleic acid formation. Alfalfa hydroperoxide lyase catalyzed n-hexanal formation from 13-hydroperoxylinoleic acid, but cucumber one catalyzed formation of n-hexanal and trans-2-nonenal from 13- and 9-hydroperoxylinoleic acid, respectively.

7. From the above results, the biosynthetic pathway for C6-aldehyde formation in etiolated alfalfa and cucumber seedlings is established that C6-aldehydes (n-hexanal, cis-3-hexenal and trans-2-hexenal) are produced from linoleic acid and linolenic acid via their 13-hydroperoxides by lipoxygenase and hydroperoxide lyase.  相似文献   

4.
C6-aldehydes are synthesized via lipoxygenase/hydroperoxide lyase action on polyunsaturated fatty acid (PUFA) substrates in plant leaves. The source pools and subcellular location of the processes are unknown. A close relationship is found between the composition of PUFA and the composition of C6-aldehydes. In the current study, this relationship was tested using the Arabidopsis PUFA mutant lines act1, fad2, fad3, fad5, fad6, and fad7. The results indicate that C6-aldehyde formation is influenced by the alteration of C18 PUFA levels. Mutants act1 and fad5, which are deficient in C16 unsaturated fatty acids, had wild-type levels of C6-aldehyde production. Mutants deficient in the chloroplast hexadecenoic acid/oleic acid desaturase (fad6) or hexadecadienoic acid/linoleic acid desaturase (fad7) had altered C6-aldehyde formation in a pattern similar to the changes in the PUFA. Mutations that impair phosphatidylcholine desaturase activity, such as fad2 and fad3, however, resulted in increased E-2-hexenal formation. The enzymes involved in C6-aldehyde production were partially characterized, including measurement of pH optima. The differences in C6-aldehyde formation among the fatty acid mutants of Arabidopsis appeared not to result from alteration of lipoxygenase/hydroperoxide lyase pathway enzymes. Investigation of the fatty acid composition in leaf phospholipids, glycolipids, and neutral lipids and analysis of the fatty acid composition of chloroplast and extrachloroplast lipids indicate that chloroplasts and glycolipids of chloroplasts may be the source or major source of C6-aldehyde formation in Arabidopsis leaves.  相似文献   

5.
Isolated tea chloroplasts utilized linoleic acid, linolenicacid and their 13-hydroperoxides as substrates for volatileC6-aldehyde formation. Optimal pH values for oxygen uptake,hydroperoxide lyase and the overall reaction from C18-fattyacids to C6-aldehydes were 6.3, 7.0 and 6.3, respectively. Methyllinoleate, linoleyl alcohol and -linolenic acid were poor substratesfor the overall reaction, but linoleic and linolenic acids weregood substrates. The 13-hydroperoxides of the above fatty acidsand alcohol also showed substrate specificity similar to thatof fatty acids. Oxygen uptakes (relative Vmax) with methyl linoleate,linoleyl alcohol, linolenic acid, -linolenic acid and arachidonicacid were comparable to or higher than that with linoleic acid.In winter leaves, the activity for C6-aldehyde formation fromC18-fatty acids was raduced to almost zero. This was due tothe reduction in oxygenation. The findings presented here provideevidence for the involvement of lipoxygenase and hydroperoxidelyase in C6-aldehyde formation in isolated chloroplasts. (Received July 11, 1981; Accepted November 5, 1981)  相似文献   

6.
Kidney bean plants (Phaseolus vulgaris) were found to have thecapability to produce C6-aldehydes (hexanal and hexenals) fromlinoleic and linolenic acids. The various organs tested hadlipoxygenase and hydroperoxide lyase activities responsiblefor the C6-aldehyde formation. Young leaves showed relativelyhigh activities for C6-aldehyde formation. However, the activitiesof the leaves decreased gradually with leaf expansion. Seedlingsand seeds containing cotyledons showed low activities for C6-aldehydeformation, because of the occurrence of an inhibitory factorin the cotyledons. The substrate specificity of the enzymeswas essentially the same among the various developmental stagesof leaves tested. (Received February 5, 1982; Accepted March 19, 1982)  相似文献   

7.
When tea leaves were homogenized and incubated, the volatileC6-compounds hexanal, cis-3-hexenal, cis-3-hexenol and trans-2-hexenalwere formed much more by summer leaves than by winter leavesof tea plants (Camellia sinensis). The enzymes lipolytic acylhydrolase (LAH), lipoxygenase, fatty acid hydroperoxide lyase(HPO lyase) and alcohol dehydrogenase (ADH) and an isomerizationfactor were responsible for the sequential reactions of C6-compoundformation from linoleic and linolenic acids in tea leaf lipids,and there were seasonal changes in their activities. The tealeaf enzymes were of 3 types: LAH and lipoxygenase, which hadhigh activities in summer leaves and low activities in winterleaves; ADH, which had low activity in summer leaves and highactivity in winter ones; and HPO lyase and the isomerizationfactor, which did not seem to have any effect on the rate ofC6-compound formation throughout the year. Changes in enzymeactivities were induced by shifts in the environmental air temperaturerather than by the age of the leaves. The combined activitiesof these enzymes determined the amounts and compositions ofthe volatile C6-compounds formed, which are the factors thatcontrol the quality of the raw leaves processed for green tea. (Received October 6, 1983; Accepted December 20, 1983)  相似文献   

8.
An enzyme has been isolated from blue-green algae Oscillatoria sp. which utilizes the product, 13-hydroperoxy-9, 11-octadecadienoic acid (13-HPOD), of lipoxygenase for its substrate. This enzyme, termed hydroperoxide lyase, converts the conjugated diene 13-hydroperoxide of linoleic acid to 13-oxotrideca-9, 11-dienoic acid. The structure of the latter has been determined by ultraviolet spectroscopy and mass spectrometry. 9-HPOD is not a substrate for this enzyme. The hydroperoxide lyase from Oscillatoria sp. has a maximum of activity at pH 6.4 and 30°C. The molecular weight of the enzyme was estimated at 56,000. The enzyme was not inhibited by BW 755C, but was inhibited by molecules containing more than one hydroxyl group. Quercetin was found to be the best inhibitor of the enzyme activity. The purified hydroperoxide lyase from Oscillatoria sp. showed an apparent Km of 7.4 micromolar and a Vmax of 35 nanomoles per minute per milligram of protein for 13-HPOD. An enzymatic pathway for the biogenesis of oxodienoic acid from linoleic acid is proposed. This involves the sequential activity of lipoxygenase and hydroperoxide lyase enzymes.  相似文献   

9.
Changes in volatile content, as well as associated gene expression and enzyme activity in developing cucumber fruits were investigated in two Cucumis sativus L. lines (No. 26 and No. 14) that differ significantly in fruit flavor. Total volatile, six-carbon (C6) aldehyde, linolenic and linoleic acid content were higher during the early stages, whereas the nine-carbon (C9) aldehyde content was higher during the latter stages in both lines. Expression of C. sativus hydroperoxide lyase (CsHPL) mirrored 13-hydroperoxide lyase (13-HPL) enzyme activity in variety No. 26, whereas CsHPL expression was correlated with 9-hydroperoxide lyase (9-HPL) enzyme activity in cultivar No. 14. 13-HPL activity decreased significantly, while LOX (lipoxygenase) and 9-HPL activity increased along with fruit ripening in both lines, which accounted for the higher C6 and C9 aldehyde content at 0-6 day post anthesis (dpa) and 9-12 dpa, respectively. Volatile compounds from fruits at five developmental stages were analyzed by principal component analysis (PCA), and heatmaps of volatile content, gene expression and enzyme activity were constructed.  相似文献   

10.
Fatty acid hydroperoxide lyase (HPO lyase) was found in green and non-green tobacco cells cultured in vitro. The HPO lyase activity in non-green cells was 13-12 of that in green cells. When the cells were transferred from the light to dark conditions or vice versa, cells turned non-green or green according to the light conditions. The HPO lyase activity also changed according to the light conditions, but the changes in HPO lyase activities were not proportional to the changes in chlorophyll contents. These results suggest that at least two types of HPO lyases are present in the green cells. One type of HPO lyase is perhaps common both to the green and non-green cells; another one is chloroplastic. The fatty acid compositions of cells and substrate specificities of HPO lyase differed between green and non-green cells.  相似文献   

11.
A Zea mays callus culture containing chlorophyll was established and grown photomixotrophically. Cell chloroplast structure, and pigment and soluble protein contents were examined. Expression of some key enzymes of C4 carbon metabolism was compared with that of etiolated (heterotrophic) and green photoautotrophic leaves. Chlorophyll content of the callus was 15–20% that of green leaves. Soluble protein content of callus was half that of leaf cells. Electron microscopic observations showed that green callus cells contained only typical granal chloroplasts. Ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco, EC 4.1.1.38) activities in green callus were ca 30% those of green leaves but 2–3 times higher than in etiolated leaves. Quantitative enzyme protein determination, using antibodies specific to maize leaf Rubisco showed that the chloroplastic carboxylase represented about 7% of total soluble protein in green callus, in parallel to its low chlorophyll content. The specific activity of Rubisco in callus and leaves was unchanged. Phosphoenolpyruvate carboxylase (PEPC, EC 4.1.1.31) activity in green callus was about 20% that of green leaves and similar to that measured in etiolated leaves. Apparent Km (PEP) values (0.08 mM) for PEPC isolated from green callus and etiolated leaves were very different from values (0.5 mM) obtained with PEPC from green leaves. These kinetic characteristics together with the absence of inhibition by malate and activation by glucose-6-phosphate suggest that the properties of PEPC isolated from green callus and etiolated maize leaves are very similar to those of PEPPC from C3 plants. Using PEPC antibodies specific to green maize leaf enzyme, immunotitration of PEPC preparations containing identical enzyme units allowed complete precipitation of the green leaf enzyme with increasing antibody volumes. In contrast, 60–70% of the activity of PEPC from etiolated and green callus was inhibited, suggesting low affinity for the maize green leaf PEPC antiserum (typical C4 form). Ouchterlony double diffusion tests revealed only partial recognition of PEPC in green callus and etiolated leaves. NAD-malate dehydrogenase (NAD-MDH, EC 1.1.1.37) activity in callus was 2 and 3 times higher, respectively, than in etiolated and green leaves. NADP-malic enzyme (NADP-ME, EC 1.1.1.40) activity in callus cultures was much lower than in green leaves. All our data support the hypothesis that cultures of fully dedifferentiated chlorophyllous tissues of Zea mays possess a C3-like metabolism.  相似文献   

12.
Bioinformatic analysis and site-directed mutagenesis allowed identification of the determinants of catalysis for CYP74, which are located in the central part of the I-helix and ERR triad. Mutations K302S and T366Y in tomato allene oxide syntase LeAOS3 induced possession of hydroperoxide lyase activity. In contrast to the wild-type MtHPL enzyme that produces C12-aldoacid, mutant forms F284I, F287V, G288I, N285A, and N285T of alfalfa hydroperoxide lyase MtHPL synthesized C13- and C11-fragments. Our data provide evidence that the CYP74 family originated from a common ancestor with hydroperoxide lyase activity.  相似文献   

13.
To assess the role of lipoxygenase (LOX; EC 1.13.11.12) in plants, we increased the expression of LOX in the tissues of Nicotiana tabacum L. cv. KY 14 by over-expression of the LOX2 gene from the soybean (Glycine max (L.) Merrill) embryo. The LOX2 cDNA was manipulated by replacing its 5-untranslated sequence with the translational enhancer of the alfalfa mosaic virus (AMV), and subcloned into a plant expression vector, 3 to a duplicated cauliflower mosaic virus 35S promoter. The AMV-LOX2 construct was transferred into tobacco using Agrobacterium tumefaciens strain A281. The LOX2 was expressed in transgenic tobacco calli, leaves of transgenic plants, and their seed progeny at levels up to 0.1–0.2% of the total extracted protein. The introduced LOX2 affected fatty-acid oxidative metabolism as evidenced by a 50–529% increase in C6-aldehyde production. The impact on C6-aldehyde formation was greater than the effect on production of fatty-acid hydroperoxides. This is consistent with other studies indicating the greater propensity of soybean embryo LOX2 in generating C6-aldehydes than that of other well-characterized LOX isozymes.Abbreviations AMV alfalfa mosaic virus - CaMV cauliflower mosaic virus - IEF isoelectric focusing - kDa kilodalton - LOX lipoxygenase - SDS-PAGE sodium dodecyl sulfate-polyacrylamide gel electrophoresis We thank Bernard Axelrod (Purdue University) for supplying the lipoxygenase 2 cDNA, and Arthur G. Hunt (University of Kentucky) for supplying the pKYLX712 and pBS/AMV. The advice of Arthur G. Hunt, Chris L. Schardl, Sadik Tuzun and Dwight Tomes is greatly appreciated, as is the technical assistance of Udaya Chand and Robert Versluys.  相似文献   

14.
Cell wall preparations (elicitors) from Phytophthora megasperma var. sojae increase C2H4 formation, phenylalanine ammonia lyase activity, and glyceollin accumulation in soybean cotyledons within about 1.5, 3, and 6 hours after treatment, respectively. The immediate precursor of C2H4, 1-aminocyclopropane-1-carboxylic acid, stimulates C2H4 formation like the elicitor within 1.5 hours after administration, whereas phenylalanine ammonia lyase activity and glyceollin concentration remain unchanged. Aminoethoxyvinylglycine, a specific inhibitor of C2H4 formation in higher plants, inhibits elicitor-induced C2H4 formation by about 95% but has no effects on phenylalanine ammonia lyase or glyceollin accumulation. It was concluded that C2H4 is a signal accompanying the specific recognition process which finally leads to the induction of phytoalexin formation, but it is not functioning as a link or messenger in the induction sequence of glyceollin accumulation.  相似文献   

15.
Linolenic acid-[1-14C] was converted to 12-oxo-trans-10-dodecenoic acid, via 12-oxo-cis-9-dodecenoic acid by incubation with chloroplasts of Thea sinensis leaves. Thus, it was confirmed that linolenic acid is split into a C12-oxo-acid, 12-oxo-trans-10-dodecenoic acid, and a C6-aldehyde, trans-2-hexenal, leaf aldehyde, by an enzyme system in chloroplasts of tea leaves.  相似文献   

16.
The effect of atmospheric methyl jasmonate on the oxylipin pathway was investigated in leaves of tobacco (Nicotiana tabacum L.), cucumber (Cucumis sativa L.), and Arabidopsis thaliana (L.). Differential sensitivities of test plants to methyl jasmonate were observed. Thus, different concentrations of methyl jasmonate were required for induction of changes in the oxylipin pathway. Arabidopsis was the least and cucumber the most sensitive to methyl jasmonate. Methyl jasmonate induced the accumulation of lipoxygenase protein and a corresponding increase in extractable lipoxygenase activity. Atmospheric methyl jasmonate additionally induced hydroperoxide lyase activity and the enhanced production of several volatile six-carbon products. It is interesting that lipid hydroperoxidase activity, which is a measure of hydroperoxide lyase plus allene oxide synthase plus possibly other lipid hydroperoxide-metabolizing activities, was not changed by methyl jasmonate treatment. Methyl jasmonate selectively altered the activity of certain enzymes of the oxylipin pathway (lipoxygenase and hydroperoxide lyase) and increased the potential of leaves for greatly enhanced six-carbon-volatile production.  相似文献   

17.
As is the case with spinach ribulose bisphosphate carboxylase/oxygenase (Rubisco), [14C]carboxyarabinitol bisphosphate (CABP) bound to purified Chlorella Rubisco with a molar ratio of unity to large subunit of the enzyme. The concentration of binding sites in extracts of photosynthetic organisms was determined by reacting the extracts with [14C]-carboxypentitol bisphosphate (CPBP) and precipitating the resultant Rubisco-[14C]CABP complex with a combination of polyethylene glycol-4000 and MgCl2. Plots of the relationship between concentrations of [14C] CPBP in the reaction mixture and the precipitated [14C]CPBP gave a straight line and the concentration of binding sites were estimated by extrapolation to zero [14C]CPBP since the dissociation constant of CABP with Rubisco is 10−11 molar. Spinach, pea, and soybean leaves contained 6.4 to 6.8 milligrams Rubisco per milligram chlorophyll, corresponding to 92 to 97 ribulose bisphosphate-binding sites per milligram chlorophyll. The Rubisco content of sunflower and wheat leaves was 5.3 to 5.5 milligrams per milligram chlorophyll. The concentrations in C4 plants were not uniform and corn and Panicum miliaceum leaves contained 3 and 7 milligrams Rubisco per milligram chlorophyll. The Rubisco content of green algae was one-fifth to one-sixth that of C3 plant leaves and was affected by the CO2 concentration during growth. The content of Euglena and blue-green algae is also reported.  相似文献   

18.
The metabolism of 13-hydroperoxylinolenic acid was examined in protoplasts and homogenates prepared from mature leaves of spinach (Spinacia oleracea L.). Chloroplast membranes were the principal site for metabolism of the compound by at least two highly hydrophobic enzyme systems, hydroperoxide lyase and hydroperoxide dehydrase, the new name for an enzyme system formerly known as hydroperoxide isomerase and hydroperoxide cyclase. Hydroperoxide lyase was most active above pH 7 and could be separated from hydroperoxide dehydrase by anion exchange chromatography. Hydroperoxide dehydrase, measured by the formation of both α-ketol product and 12-oxo-phytodienoic acid, had its optimum activity in the range of pH 5 to 7. Lyase was more active than dehydrase activity when the enzymes were extracted by homogenization. The reverse was true when the enzyme activities were measured in protoplasts, which are isolated by gentle extraction methods. The variation in enzyme activity ratios with extraction methods suggests that hydroperoxide lyase is activated by plant injury and thus may function in a wound response. In the absence of injury, the normal pathway of fatty acid hydroperoxide metabolism is probably by hydroperoxide dehydrase activity. The molecular weights of both the lyase and dehydrase were approximately 220,000, as estimated by gel filtration.  相似文献   

19.
Plants produce short‐chain aldehydes and hydroxy fatty acids, which are important industrial materials, through the lipoxygenase pathway. Based on the information that lipoxygenase activity is up‐regulated in tobacco leaves upon infection with tobacco mosaic virus (TMV), we introduced a melon hydroperoxide lyase (CmHPL) gene, a tomato peroxygenase (SlPXG) gene and a potato epoxide hydrolase (StEH) into tobacco leaves using a TMV‐based viral vector system to afford aldehyde and hydroxy fatty acid production. Ten days after infiltration, tobacco leaves infiltrated with CmHPL displayed high enzyme activities of 9‐LOX and 9‐HPL, which could efficiently transform linoleic acid into C9 aldehydes. Protein extracts prepared from 1 g of CmHPL‐infiltrated tobacco leaves (fresh weight) in combination with protein extracts prepared from 1 g of control vector‐infiltrated tobacco leaves (as an additional 9‐LOX source) produced 758 ± 75 μg total C9 aldehydes in 30 min. The yield of C9 aldehydes from linoleic acid was 60%. Besides, leaves infiltrated with SlPXG and StEH showed considerable enzyme activities of 9‐LOX/PXG and 9‐LOX/EH, respectively, enabling the production of 9,12,13‐trihydroxy‐10(E)‐octadecenoic acid from linoleic acid. Protein extracts prepared from 1 g of SlPXGinfiltrated tobacco leaves (fresh weight) in combination with protein extracts prepared from 1 g of StEH‐infiltrated tobacco leaves produced 1738 ± 27 μg total 9,12,13‐trihydroxy‐10(E)‐octadecenoic acid isomers in 30 min. The yield of trihydroxyoctadecenoic acids from linoleic acid was 58%. C9 aldehydes and trihydroxy fatty acids could likely be produced on a larger scale using this expression system with many advantages including easy handling, time‐saving and low production cost.  相似文献   

20.
Peanut,Arachis hypogaea, plants were treated in the field with the bioregulator BAS 105 00W, 4-chloro-5-dimethylamino-2-phenylpyridazin-3-one, a substituted pyridazinone, at different times of development. The seeds were harvested, dried, hand-shelled, and analyzed for lipoxygenase activity and conjugated diene hydroperoxide content. Reduced lipoxygenase activity occurred when the bioregulator was applied to the plants at flowering and pegging. The conjugated diene hydroperoxide content decreased the most in peanuts when the bioregulator was applied at pegging. The apparent Km for lipoxygenase of treated peanuts with linoleic acid as substrate was the same as that for untreated peanuts.  相似文献   

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