Specific Activity Of Phenylalanine Ammonia-lyase Research Articles

The co-occurrence of drought and Fusarium solani f. sp. Phaseoli Fs4 infection exacerbates the Fusarium root rot symptoms in common bean (Phaseolus vulgaris L.)

At this pace, global warming is expected to exacerbate drought spells now and forever. Being an integral part of the plant disease triangle, the effect drought might exerts on plant-pathogen interactions is a major subject of debates nowadays. The present study examined at how severe artificially-induced severe drought could affect the common bean and the root rot causing fungus; Fusarium solani Fs4 (F. solani Fs4) pathosystem at the morphological, physiological and biochemical levels. The results indicated significantly increased seedling susceptibility (2.08-folds; incidence = 61.8%) to fusarium root rot (FRR) upon severe drought imposition. It to be drought, F. solani Fs4-infection or their combined effects, 1.5 to 2-folds seedlings growth reduction were witnessed, with altered root growth and architecture. Further, as referred to their individual counterparts, the co-occurrence of both the stressors [F. solani + Drought] resulted in 1.23 and 1.5-fold drop in the specific activity of phenyl-alanine ammonia-lyase (PAL) and polyphenol oxidase (PPO), respectively. Meanwhile, the associative effect of the two stressors culminated at 2.3, 2.5 and 1.62-folds increase in the specific activities of peroxidases (POX), catalase (CAT), and proline content, respectively. The principal component analysis (PCA) and the Pearson correlation disclosed a negative and significant correlation between the tested defense-related enzymes PAL, PPO) and the biosynthesis of the studied de-stressing enzymes (SOD, APX, and CAT) and the osmo-protectants (proline, soluble protein). These outcomes strengthened the hypothesis of a repressive crosstalk of the Abscisic acid (ABA), the major abiotic stress mediator, on the expression of the defense-related phytohormones (Jasmonic acid and ethylene).

Effect of co-treatment of microwave and exogenous l-phenylalanine on the enrichment of flavonoids in Tartary buckwheat sprouts.

Tartary buckwheat is rich in flavonoids. The application of physical processing technology and exogenous materials treatment can effectively promote grain germination and the accumulation of bioactive secondary metabolites. The content of four flavonoids, the activities of key enzymes (phenylalanine ammonia-lyase (PAL), chalcone isomerase (CHI), flavonol synthase (FLS)) and the expression of key enzyme genes (FtPAL, FtCHI, FtFLS1, FtFLS2) in Tartary buckwheat sprouts treated with microwave and l-phenylalanine (l-Phe) were investigated, and the relationship between them was analyzed to explore the mechanism of promoting flavonoid accumulation, and to provide a theoretical basis for the development of functional Tartary buckwheat sprout food. Germination can promote the synthesis of flavonoids. The contents of chlorogenic acid and rutin in 7-day sprouts increased by 13 420.63% and 225.12% compared with seeds, respectively. Under the best treatment condition T3 (microwave 250 W, 90 s, 2.9 mmol L-1 L-Phe), the specific activities of PAL, CHI and FLS in 5-day-old sprouts increased by 47.84%, 53.04% and 28.02% compared with control check (CK), respectively; and the expression of FtPAL, FtCHI and FtFlS1 increased by 39.84%, 24.78% and 33.72% compared with CK, respectively. Correlation analysis showed that the content of flavonoids in Tartary buckwheat sprouts was significantly positively correlated with the specific activities of key enzymes (P < 0.01) and dynamically correlated with genes related to the synthesis of three enzymes. It suggested that microwave and l-Phe treatment may promote the synthesis of flavonoids by promoting the expression of key enzymes genes in phenylpropane metabolism and controlling the activity of key enzymes in phenylpropane metabolism. © 2022 Society of Chemical Industry.

IMPACT OF DROUGHT STRESS ON SOME GROWTH, BIOCHEMICAL AND ANATOMICAL PARAMETERS OF Thymus vulgaris L.

Two pot experiments were conducted on the 17th and 11th of march during 2015 and 2016 growth seasons respectively at the greenhouse, Dept. Agric. Botany, Faculty of Agriculture, Ain Shams University, Shoubra El-Kheima, Egypt to investigate the impact of drought stress on some growth, biochemical and anatomical parameters of Thymus vulgaris L. Plants were exposed to two different irrigation levels: 70-80% and 30-40% of water holding capacity (WHC) as well-irrigated and drought stressed plants respectively. The results indicated that all investigated growth parameters and leaf photosynthetic pigments were decreased significantly by exposing to drought stress. There were significant increase in the concentration of reactive oxygen species (ROS) as indicated by measuring of H2O2 and malondialdehyde (MDA). A similar trend was observed in respect to osmolytes including proline, total soluble sugars and free amino acids. Also, drought stress increased significantly total soluble phenols and the specific activity of Phenylalanine ammonia-lyase (PAL) and Polyphenol oxidase (PPO), whereas, no significant differences were detected in peroxidase (POD). The anatomical examination showed that there were several significant changes associated with water limited supply in the stem and leaf of plants.

Evaluation of Borage Extracts As Potential Biostimulant Using a Phenomic, Agronomic, Physiological, and Biochemical Approach.

Biostimulants are substances able to improve water and nutrient use efficiency and counteract stress factors by enhancing primary and secondary metabolism. Premise of the work was to exploit raw extracts from leaves (LE) or flowers (FE) of Borago officinalis L., to enhance yield and quality of Lactuca sativa ‘Longifolia,’ and to set up a protocol to assess their effects. To this aim, an integrated study on agronomic, physiological and biochemical aspects, including also a phenomic approach, has been adopted. Extracts were diluted to 1 or 10 mL L–1, sprayed onto lettuce plants at the middle of the growing cycle and 1 day before harvest. Control plants were treated with water. Non-destructive analyses were conducted to assess the effect of extracts on biomass with an innovative imaging technique, and on leaf photosynthetic efficiency (chlorophyll a fluorescence and leaf gas exchanges). At harvest, the levels of ethylene, photosynthetic pigments, nitrate, and primary (sucrose and total sugars) and secondary (total phenols and flavonoids) metabolites, including the activity and levels of phenylalanine ammonia lyase (PAL) were assessed. Moreover, a preliminary study of the effects during postharvest was performed. Borage extracts enhanced the primary metabolism by increasing leaf pigments and photosynthetic activity. Plant fresh weight increased upon treatments with 10 mL L–1 doses, as correctly estimated by multi-view angles images. Chlorophyll a fluorescence data showed that FEs were able to increase the number of active reaction centers per cross section; a similar trend was observed for the performance index. Ethylene was three-fold lower in FEs treatments. Nitrate and sugar levels did not change in response to the different treatments. Total flavonoids and phenols, as well as the total protein levels, the in vitro PAL specific activity, and the levels of PAL-like polypeptides were increased by all borage extracts, with particular regard to FEs. FEs also proved efficient in preventing degradation and inducing an increase in photosynthetic pigments during storage. In conclusion, borage extracts, with particular regard to the flower ones, appear to indeed exert biostimulant effects on lettuce; future work will be required to further investigate on their efficacy in different conditions and/or species.

A kinetic model for flavonoid production in tea cell culture.

As one of the strategies for efficient production of a metabolite from cell cultures, a kinetic model is very useful tool to predict productivity under various culture conditions. In this study, we propose a kinetic model for flavonoid production in tea cell culture based on the cell life cycle and expression of PAL, the gene encoding phenylalanine ammonia-lyase (PAL)-the key enzyme in flavonoid biosynthesis. The flavonoid production rate was considered to be related to the amount of active PAL. Synthesis of PAL was modelled based on a general gene expression/translation mechanism, including the transcription of DNA encoding PAL into mRNA and the translation of PAL mRNA into the PAL protein. The transcription of DNA was assumed to be promoted at high light intensity and suppressed by a feedback regulatory mechanism at high flavonoid concentrations. In the model, mRNA and PAL were considered to self-decompose and to be lost by cell rupture. The model constants were estimated by fitting the experimental results obtained from tea cell cultures under various light intensities. The model accurately described the kinetic behaviors of dry and fresh cell concentrations, glucose concentration, cell viability, PAL specific activity, and flavonoid content under a wide range of light intensities. The model simulated flavonoid productivity per medium under various culture conditions. Therefore, this model will be useful to predict optimum culture conditions for maximum flavonoid productivity in cultured tea cells.

Correlation of Phenylalanine ammonia lyase (PAL) and Tyrosine ammonia lyase (TAL) activities to phenolics and curcuminoid content in ginger and its wild congener, Zingiber zerumbet following Pythium myriotylum infection

The biochemical basis of resistance exhibited by a wild Zingiber species, Zingiber zerumbet (L.) Smith, towards the economically devastating soft-rot disease caused by necrotrophic Pythium myriotylum was investigated. Quantification of phenolic compounds revealed higher total phenolic (TP), total flavonoid (TF) and total tannin (TT) content in the uninfected susceptible ginger (Z. officinale) cultivar compared to the resistant taxon. However systemic induction in activities of rate-limiting enzymes of phenolic biosynthetic pathway, phenylalanine ammonia lyase (PAL) and tyrosine ammonia lyase (TAL), were observed in the resistant wild taxon. In the ginger cultivar, even though the inherent PAL specific activity was observed to be higher (24.2 ± 1.9 U mg−1) compared to the wild taxon (4.2 ± 0.8 U mg−1), a subsequent gradual decrease in both PAL and TAL activities were observed following infection of rhizomes with P. myriotylum. This was in contrast to the gradual increase in PAL (13.1 ± 0.8 U mg−1) and TAL (442.5 ± 35.1 U mg−1) specific activity after 5 days post infection (dpi) in the wild taxon. Subsequent HPLC analysis of rhizomes showed an increase in total curcuminoid content in the wild taxon compared to the ginger cultivar. Results are indicative of phenylpropanoid pathway regulation in a manner such that the induced defense metabolites contribute to restrict pathogen invasion in the resistant wild taxon.

Participation of phenylalanine ammonia-lyase (PAL) in increased phenolic compounds in fresh cold stressed walnut (Juglans regia L.) kernels

Mature fresh walnut (Juglans regia L.) kernels showed increased phenolics under cold storage in a previous study. The aim of the present work was to investigate changes in individual soluble phenolic compounds under cold stress in relation to changes in total and specific activity of phenylalanine ammonia-lyase (PAL) as an approach to study the association of particular compounds with phenylpropanoid pathway (PhP). Treatments with inhibitors of phenylalanine ammonia-lyase (PAL), mRNA, RNA and protein synthesis, as well as, activities of the enzymes polyphenol oxidase (PPO) and peroxidase (POD) were included in the experiment. Treated samples were stored at 1°C and 90% RH for 20d. Results revealed increases in specific and total activity of PAL during cold storage. Among phenolics, 2,4-dihydroxybenzoic acid and protocatechuic acid ethyl ester were identified by HPLC–DAD–ESI-MS. Increases in acids 4-hydroxybenzoic, 2,4-dihydroxybenzoic, syringic and vanillic agreed with PAL activity increases, whereas in ellagic were largely independent on PAL. Increases in protocatechuic acid and decreases in protocatechuic acid ethyl ester exhibited no direct relation to PAL.

Diadenosine polyphosphates (Ap3A and Ap4A) behave as alarmones triggering the synthesis of enzymes of the phenylpropanoid pathway in Arabidopsis thaliana

It is known that cells under stress accumulate various dinucleoside polyphosphates, compounds suggested to function as alarmones. In plants, the phenylpropanoid pathways yield metabolites protecting these organisms against various types of stress. Observations reported in this communication link these two phenomena and provide an example of a metabolic “addressee” for an “alarm” signaled by diadenosine triphosphate (Ap3A) or diadenosine tetraphosphate (Ap4A). In response to added Ap3A or Ap4A, seedlings of Arabidopsis thaliana incubated in full nutrition medium increased both the expression of the genes for and the specific activity of phenylalanine ammonia-lyase and 4-coumarate:coenzyme A ligase, enzymes that control the beginning of the phenylpropanoid pathway. Neither adenine mononucleotides (AMP, ADP or ATP) nor adenosine evoked such effects. Reactions catalyzed in vitro by these enzymes were not affected by Ap3A or Ap4A.

Activity of enzymes associated with the enzymatic browning of minimally processed potatoes

The purpose of the present study was to evaluate the effect of different potato cultivars and storage temperatures on the specific activity of phenylalanine ammonia-lyase (PAL), polyphenol oxidase (PPO) and peroxidase (POD) in minimally processed potatoes. Potato cultivars Agata, Asterix and Monalisa were selected, washed, peeled, diced, sanitized, centrifuged, vacuum- packed and stored at 5 and 15°C for 9 and 5 days, respectively. There was an increase in the enzymatic activity in all the cultivars stored at 15°C. The cultivars 'Agata' and 'Asterix' stored at 5ºC did not differ significantly between them for the PAL, PPO and POD activities. The PAL, PPO and POD activities were also influenced by the storage temperature. The cultivars Agata and Asterix were more suitable in minimal processing than 'Monalisa', which was more susceptible to oxidative browning.

The response of phenylalanine ammonia‐lyase, polyphenol oxidase and phenols to cold stress in the olive tree (Olea europaea L. cv. Picual)

AbstractBACKGROUND: In this study the activities of phenylalanine ammonia‐lyase (PAL) and polyphenol oxidase (PPO) and the concentrations of hydroxytyrosol, tyrosol and oleuropein in olive tree (Olea europaea L. cv. Picual) leaves were investigated before and after cold stress by freezing. The air temperature fell to below −7 °C and, according to the specific field conditions, four categories of orchard were selected: not cold stressed (NS), lightly cold stressed (LS), moderately cold stressed (MS) and heavily cold stressed (HS).RESULTS: In LS and MS samples the PAL specific activity at saturated substrate concentration rose 4.8‐ and 1.9‐fold respectively compared with NS samples. In HS samples the PAL activity declined by 47% compared with NS samples. A low level of PAL protein was detected in all samples affected by cold stress. In LS, MS and HS samples the PPO specific activity at saturated substrate concentration was 1.9‐, 4.4‐ and 9.8‐fold higher respectively than in NS samples. Km values also increased after cold stress. In MS and HS samples the concentration of oleuropein was 69 and 82% higher respectively than in NS samples, while the concentrations of hydroxytyrosol and tyrosol decreased.CONCLUSION: The response of PAL activity may be part of a recovery process of the olive leaf against cold stress, while the response of PPO and oleuropein may be part of an antioxidant protection mechanism. Copyright © 2009 Society of Chemical Industry

Phenylalanine ammonia‐lyase and phenolic compounds in leaves and fruits of Olea europaea L. cv. Picual during ripening

AbstractBACKGROUND: The kinetic and molecular properties of phenylalanine ammonia‐lyase (PAL) in leaves and fruit of the olive tree (Picual variety) have been studied during the seasonal process of fruit maturation. The concentrations of total phenolic compounds, oleuropein, hydroxytyrosol and tyrosol, have also been determined. This study has been made in rainfed 30‐year‐old olive trees in Jaén, Spain, cultivated by the traditional method.RESULTS: PAL specific activity was assayed and hyperbolic kinetics were observed in both organs. The Km value for L‐Phe was 0.22 mmol L−1 in leaf and 0.26 mmol L−1 in fruit. In leaf, the highest PAL specific activity was found in the stage prior to veraison. By immunoblot, a PAL‐immunoreactive 75 kDa polypeptide was detected in leaf and fruit. In leaf, the level of this protein progressively rose until the last stages of ripening at the same time that total phenols increased. In fruit, PAL activity and protein change as in two series coinciding with different fruit‐maturation period. By immunohistochemistry under light microscopy, PAL was located in the epidermis and parenchyma cells of leaf and fruit.CONCLUSION: These results demonstrate the involvement and regulation of PAL during fruit ripening of olive, cv. Picual. Copyright © 2008 Society of Chemical Industry

Induction of Phenylalanine ammonia-lyase and lipoxtgenase in cotton seedlings by mechanical wounding and aphid infestation*

Abstract It has been suggested that infestation of plants causes increases in the activities of phenylalanine ammonia-lyase (PAL) and lipowygenase (LOX), key enzymes in the phenolic compounds synthesis pathway and the octadecanoid pathway, respectively. The purpose of this work is to investigate whether he infestation of cotton aphid (Aphis gossypii) and mechanical wound can cause the induction of PAL and LOX activities in cotton seedlings, and whether the induction occurs in healthy seedlings growing nearby the attacked ones. The specific activities of PAL and LOX were measured using spectrophotometric method after aphid infestation and mechanical wounding. Result indicated that PAL activity and LOX activity were greatly induced by mechanical wounding and aphid infestation in cotton seedlings. The induction of PAL and LOX occured not only in wounded and infested seedlings but also in intact healthy seedlings growing nearby. After exposed to the aphid infestation-induced volatiles, the specific activity of PAL in cotton seedlings increased by 6% at 24 h, 80% at 48 h, 235% at 72 h compared to the control, and the specific activity of LOX increased by 18% at 24 h, 34% at 48 h, 24% at 72 h, 200% at 48 h, 164% at 72 h, respectively and the specific activity of LOX increased by 28% at 24 h, 37% at 48 h, 8% at 72 h, respectively. It suggests that the included volatiles are involved in plant-plant communication as airborne transferred signals.

Cadmium toxicity is reduced by nitric oxide in rice leaves

We evaluate the protective effect of nitric oxide (NO) against Cadmium (Cd) toxicity in rice leaves. Cd toxicity of rice leaves was determined by the decrease of chlorophyll and protein contents. CdCl2 treatment resulted in (1) increase in Cd content, (2) induction of Cd toxicity, (3) increase in H2O2 and malondialdehyde (MDA) contents, (4) decrease in reduced form glutathione (GSH) and ascorbic acid (ASC) contents, and (5) increase in the specific activities of antioxidant enzymes (superoxide dismutase, glutathione reductase, ascorbate peroxidase, catalase, and peroxidase). NO donors [N-tert-butyl-α-phenylnitrone, 3-morpholinosydonimine, sodium nitroprusside (SNP), and ASC + NaNO2] were effective in reducing CdCl2-induced toxicity and CdCl2-increased MDA content. SNP prevented CdCl2-induced increase in the contents of H2O2 and MDA, decrease in the contents of GSH and ASC, and increase in the specific activities of antioxidant enzymes. SNP also prevented CdCl2-induced accumulation of NH4 +, decrease in the activity of glutamine synthetase (GS), and increase in the specific activity of phenylalanine ammonia-lyase (PAL). The protective effect of SNP on CdCl2-induced toxicity, CdCl2-increased H2O2, NH4 +, and MDA contents, CdCl2-decreased GSH and ASC, CdCl2-increased specific activities of antioxidant enzymes and PAL, and CdCl2-decreased activity of GS were reversed by 2-(4-carboxy-2-phenyl)-4,4,5,5-tetramethyl-imidazoline-1-oxyl-3-oxide, a NO scavenger, suggesting that protective effect by SNP is attributable to NO released. Reduction of CdCl2-induced toxicity by NO in rice leaves is most likely mediated through its ability to scavenge active oxygen species including H2O2.

Effects of PAL-specific inhibition on suppression of activated defence and quantitative susceptibility of oats to Erysiphe graminis

Conidia of Erysiphe graminis DC f.sp. avenae ex. Merat were applied at various densities (low = approximately 30 mm −2, medium = approximately 100 mm −2, high = approximately 300–400 mm −2) to first leaves of spring oat ( Avena sativa) cvs Selma and Maldwyn. As previously published, high inoculum density results in activated (enhanced) host cell defence to fungal penetration, and this correlates with elevation in the frequency and intensity of host cell autofluorescent responses. In the current study, leaves were infused with either water (control treatment) or the phenylalanine ammonia lyase (PAL) inhibitors α-aminooxy-β-phenylpropionic acid (AOPP) or α-aminooxy acetic acid (AOA) at concentrations known to be inhibitory to PAL from oats. Results confirm and extend previous observations showing that these PAL inhibitors suppress localized autofluorescent host cell responses to fungal germ tube contact and increase susceptibility to penetration (haustorium formation) from appressoria. The PAL inhibitors, particularly AOPP, suppressed activated defence due to inoculum density and decreased the frequency and intensity of host cell autofluorescence responses. The data support the hypotheses that the activated defence is related to autofluorogenic host responses, and that the autofluorogens are phenolic compounds. Involvement of phenolic compound metabolism in activated defence was shown by assays of the specific activity of PAL from control cv. Selma leaves inoculated with low and high densities of conidia. Compared to uninoculated controls, the specific activity of PAL was enhanced at 6 h after inoculation (when responses to the primary germ tubes of E. graminis were evident), but there was little difference between low and high density inoculations at this time. At 18 h after inoculation (when responses to fungal appressoria were evident), the specific activity of PAL was substantially greater from high than from low inoculum density inoculations. In a coincidental observation, brown colouration (suggesting phenol oxidation) visible in many epidermal cell papillae associated with appressorial attack in control leaves, was not present in most papillae from AOPP-treated leaves.

Rosmarinic acid production in Coleus cell cultures

Cell suspension cultures of Coleus blumei Benth. have been found to accumulate 8-11% of their dry weight as rosmarinic acid (α-O-caffeoyl-3,4-dihydroxyphenyl-lactic acid). Actively-growing tissue converts >20% of exogenously supplied phenylalanine and tyrosine to the caffeoyl ester and this high rate of synthesis coincides with an increase in phenylalanine ammonia-lyase specific activity. Administration to the cultures of known phenylpropanoid precursors of rosmarinic acid failed to enhance the latter's production and in some cases inhibited it.

Specific effects on enzyme activities upon dilution of Petroselinum hortense cell cultures into water

Large increases in the specific activities of phenylalanine ammonia-lyase (EC 4.3.1.5) and p-coumarate:CoA ligase (EC 6.2.1.-) occurred within a few hours after dilution of cultured Petroselinum hortense cells into water. No significant changes in the total amount of extractable protein and in the activities of chalcone isomerase (EC 5.5.1.6) and glutamate dehydrogenase (EC 1.4.1.2) were observed under the same conditions. The time course for the change in phenylalanine ammonia-lyase activity included a lag period of 2–3 h, a peak about 13 h after the onset of induction, and a subsequent period of rapid decline. The inducible amount of enzyme activity was greatly dependent upon the degree of dilution of the cells into water. Simultaneous induction by dilution and irradiation of the cells with white, fluorescent light resulted in an increase in the phenylalanine ammonia-lyase level of activity which exceeded that calculated from the sum of the separately induced levels. Consecutive inductions, first by dilution and then 5 h later by irradiation, each required a lag period of 2–3 h. Actinomycin D or cycloheximide were inhibitors of the induction. While the total protein-synthesizing capacity in vitro was not significantly influenced by the dilution of cells, analysis of the labeled products on polyacrylamide gels demonstrated small but significant changes in the radioactivity profiles. The results are consistent with the hypothesis that dilution of the cells into water reduces the concentration of one or more compounds of cellular origin, thereby stimulating the rate of de novo synthesis of a limited number of proteins.

Effects of light on phenylalanine ammonia-lyase activity in dark-grown Zea mays (L.) seedlings

The effects of continuous white light on phenylalanine ammonia-lyase (PAL) activity in Zea mays L. roots and mesocotyls was studied. White light increased enzyme specific activity in both roots and mesocotyls. Anthocyanin accumulation in roots and mesocotyls in continuous white light up to 48 h correlated with the kinetics of light-induced PAL specific activity, anthocyanin biosynthesis lagging several hours behind light-induced PAL specific activity. When illumination was given for varying time periods followed by a dark period which ended 48 h after the beginning of the illumination period, kinetics of anthocyanin biosynthesis based on time in the light corresponded well with the light-induced PAL specific activity, indicating a possible causal relationship. Level of enzyme activity per unit of fresh weight was increased by light in both root and mesocotyl, however, total enzyme activity per mesocotyl was not changed by light as was total enzyme activity per root.