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1.
Chlorophyllase activities and chlorophyll degradation during leaf senescence in non-yellowing mutant and wild type of Phaseolus vulgaris L 总被引:6,自引:0,他引:6
The activities of chlorophyllase, contents of pigments including
chlorophyll a and b,
chlorophyllide a and b, and
phaeophorbide a during leaf senescence under low
oxygen (0.5% O2) and control (air) were investigated in a non-yellowing
mutant and wild-type leaves of snap beans (Phaseolus
vulgaris L.). Chlorophyllase from leaf tissues had maximum
activity when incubated at 40C in a mixture
containing 50% acetone. In both mutant and wild type, chlorophyllase
activity was the highest in freshly harvested non-senescent leaves and
decreased sharply in the course of senescence, indicating that the loss of
chlorophylls in senescing leaves is not directly related to the activity of
chlorophyllase and that chlorophyllase activity is not altered in the
mutant. The wild type had higher ratios of chlorophyll
a to chlorophyll b than the
mutant and chlorophyll a : b ratios increased during
senescence in both types. In the senescent mutant leaves, accumulations of
chlorophyllide a and chlorophyllide
b were detected, but no phaeophorbide
a was found. Chlorophyllide b had
a greater accumulation than chlorophyllide a in the
early stage of senescence. Low oxygen treatment not only delayed
chlorophyll degradation but also enhanced the accumulations of
chlorophyllide a and b and
lowered the ratios of chlorophyll a to chlorophyll
b. 相似文献
2.
B. T. Steer 《Physiologia plantarum》1970,23(3):505-512
In the presence of large accumulations of protochlorophyllide, derived from exogenous δ-aminolevulinic acid, chlorophyll synthesis in excised leaves of two varieties of barley was less than in untreated leaves. In oat leaves the accumulated protochlorophyllide, from exogenous δ-laminolevulinic acid, stimulated chlorophyll synthesis to above the control level. — These relationships could only be demonstrated when phtodestruction of pigments was minimised by the use of flash illumination (2 milliseconds every 3 minutes). — These was no evidence from in vivo absorption spectra that the pigments in the barley leaves were different to those in leaves studied by other workers. However, the presence of the accumulated protochlorophyllide appeared to prevent the shift of the chlorophyll absorption maximum from 673 nm to 677 nm. — Possible mechanisms of inhibition are discussed. 相似文献
3.
D. Todorov V. Alexieva E. Karanov D. Velichkov V. Velikova 《Journal of Plant Growth Regulation》1992,11(4):233-238
The effect of some dicarboxylic acid monoesters on growth, chlorophyll content, chlorophyllase (EC 3.1.1.14), and total peroxidase (EC 1.11.1.7.) activities was examined in detached and intact leaves of maize (Zea mays) plants grown in a greenhouse. The -monomethyl ester of itaconic acid (MEIA) at 1250 ppm had no effect on growth. However, application of the monoethyl ester of succinic (MESA) and monoethyl ester of adipic (MEAdA) acids (1250 ppm) resulted in an increased leaf area, fresh and dry weight of leaves and stems. These compounds retarded chlorophyll degradation in both detached and intact leaves. Chlorophyllase activity of the control and treated leaves was measured and related to chlorophyll content. Delaying of senescence by treatment with monoesters resulted in greater chlorophyll and protein content, compared with the control. However, the chlorophyllase activity/chlorophylla ratio in the treated plants decreased. Total peroxidase activity was higher in senescent leaves, but all treatments inhibited the increase of this enzyme activity. Prolonged carbon assimilative activity and enhanced leaf water use efficiency in treated plants was noted. 相似文献
4.
Yang Liao Kun An Xiao Zhou Wen-Jun Chen Ben-Ke Kuai 《植物学报(英文版)》2007,49(4):531-539
Chlorophyllase (EC 3.1.1.14) is involved in the first step of chlorophyll degradation. Isolation of chlorophyllase genes greatly facilitates characterization of chlorophyllase properties and elucidation of molecular regulation of their in vivo activities. There are two chlorophyllase genes, AtCLH1 and AtCLH2, in Arabidopsis thaliana. The in vivo roles of AtCLH1 have been reported previously. However, few studies have been carried out on AtCLH2. Here, we show that purified recombinant Chlase2, encoded by AtCLH2, exhibits in vitro chlorophyllase activity. Interestingly, "activation" of in vitro activity of the recombinant Chlase2 required higher concentrations of a detergent or a polar solvent. To determine its activity in vivo, the expression of AtCLH2 was inhibited by RNA interference. RNAi plants showed decreased contents of chlorophyllide without a substantial change in the total amount of the extractable chlorophyll and consequently presented lower chlorophyllide to chlorophyll ratios in their leaves. In addition, the two AtCLHs exhibited differential expression patterns. Our results suggest that AtCLH2 might play a distinctive role in chlorophyll catabolism in vivo. 相似文献
5.
Sequence of Chloroplast Degreening in Calamondin Fruit as Influenced by Ethylene and AgNO(3) 总被引:3,自引:3,他引:0 下载免费PDF全文
Purvis AC 《Plant physiology》1980,66(4):624-627
C2H4 disrupts the internal membranes of the chloroplast and induces an increase in chlorophyllase activity in degreening calamondin [x Citrofortunella mitis (Blanco) Ingram and Moore] fruit. Whether the loss of chlorophyll in the peel is causally related to breakdown of the chloroplast and/or chlorophyllase activity is not readily apparent. Chlorophyllase levels were inversely related to chlorophyll content, but electron micrographs also showed that internal membranes of the chloroplasts were disrupted simultaneously with the decrease in chlorophyll content. Silver, a potent inhibitor of C2H4-mediated effects, retarded the loss of chlorophyll in calamondin rind, reduced the C2H4-induced increase in chlorophyllase level, and prevented the disruption of the chloroplast membranes. The results do not permit the proposal of a mechanism of C2H4 metabolism in the degreening of calamondin fruit. 相似文献
6.
Abstract Chlorophyllase activity and photosynthetic membrane proteins of two lots of Hedera helix were studied. Both the HV variety with variegated leaves and HC variety with green leaves were observed. Proteins from green, white and white-green portions of leaves differing in age were compared using dodecyl-sulfate-polyacrylamide gel electrophoresis. The white or white-green regions of Hedera helix var. aureomarginata (HV) did not differ significantly in chlorophyllase activity. 相似文献
7.
Total chlorophyll content and chlorophyllase (chlorophyll-chlorophyllido hydrolase EC 3.1.1.14) activity in fresh leaves of
Piper betle L. landrace KS was, respectively, twofold higher and eight fold lower than KV, showing negative correlation between chlorophyll
and chlorophyllase activity. Specific chlorophyllase activity was nearly eightfold more in KV than KS. ORF of 918 nt was found
in cloned putative chlorophyllase cDNAs from KV and KS. The gene was present as single copy in both the landraces. The encoded
polypeptide of 306 amino acids differed only at two positions between the KV and KS; 203 (cysteine to tyrosine) and 301 (glutamine
to glycine). Difference in chlorophyllase gene expression between KV and KS was evident in fresh and excised leaves. Up regulation
of chlorophyllase gene by ABA and down regulation by BAP was observed in both the landraces; however, there was quantitative
difference between KV and KS. Data suggests that chlorophyllase in P. betle is involved in chlorophyll homeostasis and chlorophyll loss during post harvest senescence. 相似文献
8.
Abstract The effects of kinetin on chlorophyll breakdown in irradiated barley leaves. — Kinetin is shown to inhibit the breakdown of chlorophyll in isolated barley leaves. Moreover the kinetin is shown to inhibit, even if with lower effect, the chlorophyll breakdown in irradiated barley leaves. This possible correlation of these observations, with kinetin promoted protein synthesis is suggested. 相似文献
9.
Rispens T Leeuwen Av Vennegoor A Killestein J Aalberse RC Wolbink GJ Aarden LA 《Analytical biochemistry》2011,(2):271-276
Chlorophyllide a is a metabolite late in the biosynthesis of chlorophylls and bacteriochlorophylls. Isolation procedures for chlorophyllide a from Rhodobacter capsulatus CB1200 and barley (Hordeum vulgare L.) are described and compared. R. capsulatus CB1200 is a double mutant in the bacteriochlorophyllide a biosynthetic pathway, and chlorophyllide a is excreted by the cells when grown in Tween 80-containing liquid medium. It was purified by liquid or solid phase extraction, yielding 7 mg of chlorophyllide a from 1 L of culture. In a second approach, intrinsic chlorophyllase activity was used to dephytylate chlorophyll in an acetonic preparation of leaves of wild-type or chlorophyll b-deficient barley. Purification was achieved by liquid phase extraction, yielding 14 μg of chlorophyllide a per gram of barley leaves. Chlorophyllide a was identified by thin layer chromatography, absorption spectroscopy, and mass spectrometry. 相似文献
10.
Ribonuclease and Chlorophyllase Activities in Senescing Leaves 总被引:3,自引:0,他引:3
The activities of two enzymes, ribonuclease and chlorophyllase were investigated during the senescence of leaves. Ribonuclease activities were measured in primary leaves of Phaseolus vulgaris, and related to the levels of nucleic acid, protein and chlorophyll. Similarly, changes in chlorophyllase activity during senescence of leaves of Raphanus sativus were measured and related to chlorophyll. During senescence the levels of each enzyme as well as its respective substrate declined. Retardation of senescence, by excision of young tissue from intact plants or by treatment of detached leaves with cytokinins resulted in a maintainace of both the substrate and enzyme levels. It was concluded that high levels of ribonuclease and chlorophyllase activity are not linked directly with the degradation of RNA and chlorophyll during leaf senescence. 相似文献
11.
Chlorophyllase (chlorophyll-chlorophyllidohydrolase, EC 3.1.1.14) was isolated and purified from Phaseolus vulgaris L. chloroplasts and etioplasts dissolved in 1% Triton X-100 and 10% glycerol. A 100 and 40-fold purification, respectively, was achieved. Enzyme preparations from both sources had similar affinities for chlorophyll a when assayed in a Triton X-100 medium. When electrophoresed in sodium dodecyl sulphate polyacrylamide gels the major band in both preparations migrated as a peptide of 30,000 daltons. Chlorophyll containing liposomes were also used as a substrate for chlorophyllase. The rate of hydrolysis did not follow Michaelis-Menten kinetics. When chlorophyllide a or methyl chlorophyllide a was incorporated in the liposomes, then in the presence of phytol dissolved in methanol, methylchlorophyllide a and chlorophyll a were shown to be synthesized. Apparently the purified enzyme in the presence of lipids, is endowed with both synthetic and hydrolytic activity.Abbreviations DEAE
diethylaminoethyl
- MeOH
methanol
- SDS
sodium dodecyl sulphate 相似文献
12.
Biogenesis of the photosynthetic apparatus in greening etiolated leaves of barley (Hordeum vulgare L) was investigated by an approach permitting investigation of this process under conditions that minimize differences in
plastid development. Distributions of barley leaves greening for 24 h as to chlorophyll content and of chloroplast grana as
to number of thylakoids were shown to be of a multimodal character. The shape of time-course curves of chlorophyll accumulation
in local sites of greening etiolated leaves was of a stepped or (at the end of greening) undulated character. The stepwise
accumulation of chlorophyll was accompanied by wave-like changes in chlorophyll b/a ratio, intensity of low-temperature chlorophyll fluorescence and photosynthetic activity with minima at the time points of
transition to accelerated chlorophyll accumulation. It is assumed that (1) development of the photosynthetic apparatus in
local sites of greening etiolated leaves occurs stepwise, from one steady level to another, but not as gradually as is generally
accepted, and (2) every separate step in development of the photosynthetic apparatus seems to begin with formation of photosystem
cores and to end with the synthesis of light-harvesting complexes.
This revised version was published online in June 2006 with corrections to the Cover Date. 相似文献
13.
Localization of chlorophyllase in the chloroplast envelope 总被引:9,自引:0,他引:9
Chlorophyllase catalyzes the first step in the catabolic pathway of chlorophyll. It is a constitutive enzyme located in chloroplast membranes. In isolated plastids the hydrolysis of the endogenous chlorophyll does not take place unless the membranes are solubilized in the presence of detergent. The structural latency of chlorophyllase activity appears to be due to the differential locations of substrate and enzyme within the plastids. Envelope membranes prepared from both chloroplasts and gerontoplasts contain chlorophyllase activity. The isolation of envelopes is associated with a marked increase in chlorophyllase activity per unit of protein. Yields of chlorophyllase and of specific envelope markers in the final preparations are similar, suggesting that the enzyme may be located in the envelope. It is hypothesized that the breakdown of chlorophyll during leaf senescence requires a mechanism that mediates the transfer of chlorophyll from the thylakoidal pigment-protein complexes to the sites of catabolic reactions in the envelope.Abbreviations ACT
acyl CoA thioesterase
- Chl
chlorophyll
- Chlide
chlorophyllide
- PC
phosphatidylcholine 相似文献
14.
Chlorophyllase (EC 3.1.1.14) is involved in the first step of chlorophyll degradation. Isolation of chlorophyllase genes greatly facilitates characterization of chlorophyllase properties and elucidation of molecular regulation of their in vivo activities. There are two chlorophyllase genes, AtCLH1 and AtCLH2, in Arabidopsis thallana. The in vivo roles of AtCLH1 have been reported previously. However, few studies have been carried out on AtCLH2. Here,we show that purified recombinant Chlase2, encoded by AtCLH2, exhibits in vitro chlorophyllase activity. Interestingly,"activation" of in vitro activity of the recombinant Chlase2 required higher concentrations of a detergent or a polar solvent. To determine its activity in vivo, the expression of AtCLH2 was inhibited by RNA interference. RNAi plants showed decreased contents of chlorophyllide without a substantial change in the total amount of the extractable chlorophyll and consequently presented lower chlorophyllide to chlorophyll ratios in their leaves. In addition, the two AtCLHs exhibited differential expression patterns. Our results suggest that AtCLH2 might play a distinctive role in chlorophyll catabolism in vivo. 相似文献
15.
《Bioscience, biotechnology, and biochemistry》2013,77(9):1660-1663
We investigated the total content of pheophorbide a (PB a), which is sum of the contents of newly produced PB a, including PB a initially present and that converted from chlorophyllide a (Chd a) by the chlorophyllase reaction during incubation, in green tea samples, and found that the total content of PB a markedly increased in both Sencha and Matcha, compared with the initially present PB a content in each. This result demonstrates that chlorophyllase activity still remains in green tea, even after processing fresh green leaves. A comparison of the total contents of PB a produced during the incubation of chlorophyll a (Chl a) with Sencha and fresh green leaf acetone powder indicates that the ratio of chlorophyllase activity in Sencha and in fresh green leaves was about 1:20. 相似文献
16.
《FEBS letters》1987,226(1):72-76
Chlorophyllase (chlorophyll chlorophyllidohydrolase, EC 3.1.1.14) catalyses the transesterification of chlorophylls with the surfactant Triton X-100, which is widely used in the preparation and study of this enzyme. The preparation and some properties of water-soluble tritonyl chlorophyllide esters are described. A mechanism for the role of Triton X-100 as an inhibitor in chlorophyllase-catalyzed hydrolysis and transesterification of chlorophylls is proposed. Bacteriochlorophyl a also has been employed as a substrate for green plant chlorophyllase. 相似文献
17.
18.
In the degradation of chlorophyll, chlorophyllase catalyzes the initial hydrolysis of the phytol moiety from the pigment. Since chlorophyll degradation is a defining feature of plant senescence, compounds inhibiting chlorophyllase activity may delay senescence, thereby improving shelf life and appearance of plant products. Here we describe the development of a 96-well plate-based purification and assay system for measuring chlorophyllase activity. Integrated lysis and immobilized metal affinity chromatography plates were used for purifying recombinant hexahistidine-tagged Triticum aestivum (wheat) chlorophyllase from Escherichia coli. Chlorophyllase assays using chlorophyll as a substrate showed that the immobilized fusion protein displayed kinetic parameters similar to those of recombinant enzyme purified by affinity chromatography; however, the need to extract reaction products from a multiwell plate limits the value of this assay for high-throughput screening applications. Replacing chlorophyll with p-nitrophenyl-ester substrates eliminates the extraction step and allows for continuous measurement of chlorophyllase activity in a multiwell plate format. Determination of steady state kinetic constants, pH rate profile, the inhibitory effects of metal ions and esterase inhibitors, and the effect of functional group-modifying reagents validated the utility of the plate-based system. The combined purification and assay system provides a convenient and rapid method for the assessment of chlorophyllase activity. 相似文献
19.
《Bioscience, biotechnology, and biochemistry》2013,77(3):433-437
Chlorophyllase from a diatom alga (Phaeodactylum tricornutum) was obtained and the partially purified extract has been further purified using preparative isoelectric focusing on a Rotofor cell. Three fractions, FI, FII, and FIII, were separated from the Rotofor cell and salt and ampholytes were removed to give fractions FI′, FII′, and FIII′, respectively. Enzyme fractions FI′, FII′, and FIII′, respectively. Enzyme fractions FI′, FII′, and FIII′ showed specific activities of 15.2 × 10?4, 226.7 ×10?4 and 33.8 × 10?4 µmol/mg protein/min, respectively. Most of the enzyme activity (84%) was in fraction FII′. The optimum pH for chlorophyllase activity was 8.0 for FI′ and 8.5 for both FII′ and FIII′. Apparent Km values for enzyme fractions FI′, FII′, and FIII′ were 2.1nM, 2.3nM, and 2.0 nM, respectively. Enzyme fractions FII′ and FIII′ showed higher chlorophyllase activity towards the partially purified chlorophyll when it was compared to that with the crude chlorophyll as well as with both chlorophylls a and b. However, the enzyme fraction FI′ had higher activity towards the crude chlorophyll when it was compared to that with both chlorophylls a and b, but with a preference for chlorophyll a over chlorophyll b. The inhibitory effect of diisopropyl flurophosphate (DIFP) on chlorophyllase activity demonstrates a noncompetitive inhibitor kinetics with Ki values of 1.29mM, 2.14mM, and 0.71mM for FI′. FII′, and FIII′, respectively. 相似文献
20.
During senescence of flowering plants, only breakdown products derived from chlorophyll a were detected although b disappears, too (Matile et al., 1996, Plant Physiol 112: 1403–1409). We investigated the possibility of chlorophyll b reduction during dark-induced senescence of barley (Hordeum vulgare L.) leaves. Plastids isolated from senescing leaves were lysed and incubated with NADPH. We found 71-hydroxy-chlorophyll a, 71-hydroxy-chlorophyllide a, and, after incubation with Zn-pheophorbide b, also Zn-71-hydroxy-pheophorbide a, indicating activity of chlorophyll(ide) b reductase. The highest activity was found at day 2 of senescence when chlorophyll breakdown reached its highest rate. Chlorophyllase
reached its highest activity under the same conditions only at days 4–6 of senescence. Based on the chlorophyll b reductase activity of plastids at day 2.5 of senescence (=100%), the bulk of activity (83%) was found in the thylakoids and
only traces (5%) in the envelope fraction. Chlorophyll b reduction is considered to be an early and obligatory step of chlorophyll b breakdown.
Received: 22 February 1999 / Accepted: 24 March 1999 相似文献