Acid Phosphatase Isozymes Research Articles

A Study on Evaluation of Alkaline and Acid Phosphatase Isozymes During Different Developmental Stages of New Breeding Lines and Races of Bombyx mori L

It is interesting to note that different silkworm races reared in laboratory offer an important testing ground for the application of biochemical methods to taxonomic problems. Moreover, there is scarcity of knowledge on enzyme studies in new breeding lines and races of silkworm specially Bombyx mori L. Therefore, we designed the present study with the main goal to evaluate the activities of alkaline and acid phosphatases quantitatively during different developmental stages of new breeding lines and races viz. Kalimpong-A (KA), B18, Pure Mysore (PM), evolved R1 and R2 of Bombyx mori L. Quantitative estimations of in alkaline and acid phosphatases were expressed in terms of enzyme activity. Alkaline and acid phosphatase activities during the different developmental stages of KA, NB18, PM, evolved R1 & R2 races were determined using Sodium-1 naphthahyl phosphate as a substrate following the dye-coupling method. The assay mixture included 2 ml of substrate and 0.2 ml of enzyme extract and incubation was made for 30 minutes at 25°c. The reaction was stopped by adding 2 ml of post coupling solution (5 parts of 4% sodium dodecyl sulphate and 2 parts of 0.2% Fast red TR salt) and colorimetric determination were made at 540 nm. Results illustrated that the activity of phosphatases was found to be different and high activity was found in the larval stage, which is feeding stage followed by pupae.

Isolation and Characterization of three Thermostable Acid Phosphatase isozymes from Cassia occidentalis (Coffee Senna) Seeds

Isolation and Characterization of three Thermostable Acid Phosphatase isozymes from Cassia occidentalis (Coffee Senna) Seeds

Effects of Specific Electric Field Stimulation on the Release and Activity of Secreted Acid Phosphatases from Leishmania tarentolae and Implications for Therapy.

Leishmaniasis is a neglected tropical disease with 1.6 million new cases reported each year. However, there are few safe, effective, and affordable treatments provided to those affected by this disease. Still under-appreciated as potential pharmaceutical targets, especially for cutaneous leishmaniasis infections, are the two isozymes of secreted acid phosphatase (SAP). These enzymes are involved in the survival of the parasite in the sand fly vector, and in infecting host macrophages. While the application of electric or electromagnetic fields as a medicinal therapeutic is not new, the utility of electric field application for the treatment of leishmaniasis is under studied. Studies involving the effects of electric fields on the cell secretion of SAP or the activity of SAP that has been secreted prior to electrical stimulation have not yet been reported. This work is the first report on the effect of specific electric fields on the activity of Leishmania tarentolae secreted acid phosphatases and the modulation of this secretion from the cells. In addition, the kinetic constants for the enzyme isoforms were determined as a function of days in culture and removal of carbohydrate from the glycosylated enzymes, while using a glycosidase, was shown to affect these kinetic constants.

English

English

Absence of an acid phosphatase isozyme locus as a marker candidate for true to typeness in woodland grape (Vitis vinifera L. ssp. sylvestris Gmelin)

<p style="text-align: justify;">The quest and conservation of existing populations of woodland grape (<em>Vitis vinifera</em> L. ssp. <em>sylvestris</em> Gmelin), the supposed progenitor of the European grapevine (<em>Vitis vinifera</em> L. ssp. <em>sativa</em>) and a significant actor in the evolution of grapevine, has great importance in preserving biodiversity. The proof of true-to-typeness is highly important in ex-situ conservation, because the contamination risk of the woodland grape populations is very high. Some characteristic “<em>sylvestris</em>” simple sequence repeats (SSR) alleles were identified, but they are only characteristic in a specific population.</p><p style="text-align: justify;">In our recent study, the SSR profiles of 32 woodland grapes were compared to those of 16 European grapevine varieties and 20 rootstocks. Morphology and SSR analyses suggested that the analysed <em>Vitis vinifera</em> ssp. <em>sylvestris</em> Gmelin accessions were true-to-type. In this report, the results of the acid phosphatase isoenzyme analyses of the same woodland grape accessions are presented and a new marker for true-to-typeness is suggested.</p>

Isozyme analysis based genetic fidelity assessment of micropropagated banana plants

Isozyme studies of micropropagated and mother plants of banana cvs. Matti, Ney Poovan, Kechulepa, Dwarf Cavendish, Malbhog, Champa, B.B. Battisa and FHIA-1 were done to test their genetic fidelity. The banding patterns as revealed by electrophoretic variations were evaluated with respect to isozymes of acid phosphatase, catalase, esterase and peroxidase as markers. The genetic fidelity of micropropagated plants and the relationship of the different cultivars were determined by dendrogram using numerical taxonomy and multivariate analysis system(NTSYS). A clustered dendrogram was prepared by unweighted pair group method using averages (UPGAMA) method. At 87% similarity, the micropropagated and mother plants were clustered in four groups reflecting their genomic constitution. Cvs. Matti (AA) and Dwarf Cavendish (AAA) with similar ‘A’ genome were categorized in Cluster I. Cluster II comprised of cvs. Ney Poovan (AB), B.B. Battisa (ABB) and FHIA-1(AAAB) with genomic constitution of both ‘A’ and ‘B’ type. Cvs. Champa (AAB) and Malbhog (AAB) with similar genome were grouped inCluster III. Cluster IV contained the cv. Kechulepa (BB) having only ‘B’ genome. However, there was no somaclonal variation among the micropropagated plants and they showed 100% genetic similarity. Thus, the isozyme studies could be a reliable marker for testing the genetic fidelity of micropropagated plants and for evaluating the diversity among the banana germplasm.

Evidence of High Gene Flow Between Samples of Horseweed (Conyza canadensis) and Hairy Fleabane (Conyza bonariensis) as Revealed by Isozyme Polymorphisms

Native polyacrylamide gel electrophoresis was used to identify polymorphisms in α- and β-esterases loci and electrophoresis in starch gel to identify polymorphism in malate dehydrogenase (MDH; EC 1.1.1.37) and acid phosphatase (ACP; EC 3.1.3.2) isozymes loci in leaf tissues from samples of horseweed and hairy fleabane populations to determine genetic diversity and population structure. Similar or differential genetic divergence between the two species may guide specific use of herbicides. For samples of plants with high genetic similarity it is possible to adopt similar mechanisms and processes for their control. The proportion of polymorphic loci was 57.14, 50.0, and 53.6%, in samples of horseweed and hairy fleabane, for EST, MDH, and ACP isozymes, respectively. A comparison of the diversity parameters in the two species showed that the number of alleles is similar in the horseweed and hairy fleabane plants. The estimated heterozygosity in horseweed and hairy fleabane was also very close. A relatively low level of population differentiation was detected between horseweed and hairy fleabane (FST= 0.0199), which suggests a substantial genetic exchange among the two species. Accordingly, estimate of gene flow was high (Nm = 12.3172) for the alleles of the lociEst,Mdh, andAcp. The Nei's identity (I) values also was high (I = 0.9561) indicating very high similarity between the twoConyzaspecies. AMOVA showed higher genetic variation within (95%) than among (5%) the two samples. The low genetic structure and high value of genetic identity was an important indication that alleles are exchanged between horseweed and hairy fleabane populations, and provides additional evidence of occurrence of outcrossing between populations or dispersion of samples of one for other site.

Molecular characterization of four morphological mutants of Neurospora crassa

Four morphological mutants (albino, ropy, conidial band and buff) of Neurospora crassa were characterized based on morphological features and molecular markers. The mycelial color of mutants buff, ropy and conidial band were more or less orange like wild parents. Albino mutant showed colorless mycelium. The germination time of conidia of wild and other mutants ranged from 5 to 7 days while in albino it was 12 days. Esterase and acid phosphatase isozymes analysis of the N. crassa mutants clearly indicated that mutation altered the carotenoid biosynthesis pathway creating the albino mutant due to the effect UV light. Most of the mutants viz; albino, conidial band and buff showed characteristic RAPD banding profile. However, no band was found in wild EmA, Ema and mutant ropy. Highest number of RAPD bands were found in albino. The mutant albino showed very different morphological and molecular features from the rest specimens. DOI: http://dx.doi.org/10.3329/dujbs.v23i1.19831 Dhaka Univ. J. Biol. Sci. 23(1): 85-91, 2014

Biochemical and molecular genetic characterization of some species of family Malvaceae, Egypt

The aim of the present study intended mainly to investigate the interrelationships between the six studied taxa namely Abutilon theophrasti, Lavatera cretica, Hibiscus trionum, Hibiscus sabdariffa, Malva parviflora and Sida alba collected from ten different accessions in Egypt belonging to family Malvaceae. Biochemical studies include protein profile using SDS-PAGE technique and three isozymes (esterase, peroxidase and acid phosphatase). The electrophoretic analysis revealed the presence of eighteen bands of molecular weight ranging from 11.3 to 115.3 KD. The highest number of bands 15 was observed in H. sabdariffa (Hs1) collected from Menia el-Kamh district and the two accessions of M. parviflora where as the lowest number 11 bands were recorded in H. trionum collected from Talkha district. Four loci of peroxidase isozyme distinguished, three loci of acid phosphatase isozyme and two loci of esterase isozyme. In regarding to random amplified polymorphic DNA technique (RAPD), ten primers were used to differentiate between these accessions. Primer OPA-4 gave the highest percentage of polymorphism (100%), while primer OP-B6 produced the lowest percentage (50%).

Physiological and Biochemical Effects of γ-Irradiation on Cowpea Plants (Vigna sinensis) under Salt Stress

Soil salinity is one of the most severe factors limiting growth and physiological response in cowpea plants. In this study, the possible role of γ-irradiation in alleviating soil salinity stress during plant growth was investigated. Increasing salinity in the soil (25, 50, 100 and 200 mM NaCl) decreased plant growth, photosynthetic pigments content, total carbohydrate content and mineral uptake compared to control, while increased total phenol content, proline, total free amino acids and lipid peroxidation. Seed irradiation with gamma rays significantly increased plant growth, photosynthetic pigments, total carbohydrate, total phenol, proline, total free amino acids and the contents of N, P, K +, Ca+2 and Mg+2 compared to non irradiated ones under salinity. On the other hand, irradiation with gamma rays decreased lipid peroxidation, Na+ and Cl- contents which may contribute in part to activate processes involved in the alleviation of the harmful effect of salt at all concentrations used (25, 50 and 100 mM) except at the high concentration (200 mM). Electrophoretic studies of α-esterase, β-esterase, polyphenol oxidase, peroxidase and acid phosphatase isozymes showed wide variations in their intensities among all treatments.

Alleviation of Cadmium Toxicity in Pisum sativum L. Seedlings by Calcium Chloride

The present investigation was carried out to study the role of calcium chloride in enhancing tolerance and reducing cadmium toxicity in pea seedlings. Some treatment with 1 and 5 mM CaCl2 mitigated cadmium stress by increasing antioxidant enzyme activities: catalase (CAT), peroxidase (POD) and polyphenol oxidase (PPO), as well as by elevating contents of ascorbic acid (ASA), tocopherol and carotenoids. On the other hand, total carbohydrate and total soluble proteins decreased with increasing cadmium concentrations in comparison with control plants. However, total phenol, total free amino acids, proline and lipid peroxidation increased with increasing concentrations of cadmium acetate. Electrophoretic studies of protein revealed that cadmium treatments alone or in combination with calcium chloride were associated with the disappearance of some bands or appearance of new bands in pea seedlings. Electrophoretic studies of α-esterase, β-esterase and acid phosphatase isozymes showed wide variations in their intensities and densities.

The secreted purple acid phosphatase isozymes AtPAP12 and AtPAP26 play a pivotal role in extracellular phosphate-scavenging by Arabidopsis thaliana

Orthophosphate (Pi) is an essential but limiting macronutrient for plant growth. Extensive soil P reserves exist in the form of organic P (Po), which is unavailable for root uptake until hydrolysed by secretory acid phosphatases (APases). The predominant purple APase (PAP) isozymes secreted by roots of Pi-deficient (–Pi) Arabidopsis thaliana were recently identified as AtPAP12 (At2g27190) and AtPAP26 (At5g34850). The present study demonstrated that exogenous Po compounds such as glycerol-3-phosphate or herring sperm DNA: (i) effectively substituted for Pi in supporting the P nutrition of Arabidopsis seedlings, and (ii) caused upregulation and secretion of AtPAP12 and AtPAP26 into the growth medium. When cultivated under –Pi conditions or supplied with Po as its sole source of P nutrition, an atpap26/atpap12 T-DNA double insertion mutant exhibited impaired growth coupled with >60 and >30% decreases in root secretory APase activity and rosette total Pi concentration, respectively. Development of the atpap12/atpap26 mutant was unaffected during growth on Pi-replete medium but was completely arrested when 7-day-old Pi-sufficient seedlings were transplanted into a –Pi, Po-containing soil mix. Both PAPs were also strongly upregulated on root surfaces and in shoot cell-wall extracts of –Pi seedlings. It is hypothesized that secreted AtPAP12 and AtPAP26 facilitate the acclimation of Arabidopsis to nutritional Pi deficiency by: (i) functioning in the rhizosphere to scavenge Pi from the soil’s accessible Po pool, while (ii) recycling Pi from endogenous phosphomonoesters that have been leaked into cell walls from the cytoplasm. Thus, AtPAP12 and AtPAP26 are promising targets for improving crop P-use efficiency.

Sustainable effects of diatomite on the growth criteria and phytochemical contents of Vicia faba plants

Recent scientific findings proved that silicon feeding will provide benefits to plant growth and yield depending on the source of Si, the mode of application , the concentration used, the plant species , the mode of action and the biotic or abiotic factor involved. To better understand the intrinsic properties of Si , this work was conducted using a series of diatomite (diatomaceous earth)at rates of 0,2.5, 5 and 10 g/kg soil, to look into the most subtle changes in the growth , yield, metabolism, electropharetic protein and isozyme pattern of faba bean ( Vicia faba) plant. Indeed, diatomite-treated plants displayed an over all better morphological increments expressed as shoot and root length , number of leaves and pods/ plant, fresh and dry weights of shoots and roots and physiological activity expressed as chlorophyll a,b and total pigments , total soluble sugars (TSS) and total sugars (TS), photosynthetic rate, leaf stomatal conductance, % of leaf relative humidity ( %LRH), net intercellular carbon dioxide (∆CO2), total nitrogen (TN) and total soluble nitrogen (TSN) and the concentrations of phosphorus (P), potassium (K), calcium(Ca) and magnesium (Mg) than non-treated faba bean plants . Furthermore, diatomites (86-89 % SiO2) induced and restored ten electrophoretic protein bands which confirmed their significance as an algal manure in accelarating the gene functions to perform, in an elevated rate, new silence genes to operate thus expressing new protein profiles. Soil fertilization by diatomites induced four esterase ( EST), three peroxidase (POD), two catalase (CAT) and three acid phosphatase (ACP) isozyme polymorphic bands which were absent in the control plants .The intensity and density of these bands were significantly increased in response to diatomite treatment. The impact of diatomite is time-dependent as it became more effective while the experiment continued and increased in a dose-related manner . Reversibly, diatomites application showed negative response in the transpiration rate and the values of both carotenoids, sodium and iron. This study highlighted the fact that diatomites are important component of the production system and should not be ignored when attempting to attribute causes for below optimum production

Biochemical and molecular characterization of AtPAP12 and AtPAP26: the predominant purple acid phosphatase isozymes secreted by phosphate‐starved Arabidopsis thaliana

Plant purple acid phosphatases (PAPs) belong to a large multigene family whose specific functions in Pi metabolism are poorly understood. Two PAP isozymes secreted by Pi-deficient (-Pi) Arabidopsis thaliana were purified from culture filtrates of -Pi suspension cells. They correspond to an AtPAP12 (At2g27190) homodimer and AtPAP26 (At5g34850) monomer composed of glycosylated 60 and 55 kDa subunit(s), respectively. Each PAP exhibited broad pH activity profiles centred at pH 5.6, and overlapping substrate specificities. Concanavalin-A chromatography resolved a pair of secreted AtPAP26 glycoforms. AtPAP26 is dual targeted during Pi stress because it is also the principal intracellular (vacuolar) PAP up-regulated by -Pi Arabidopsis. Differential glycosylation appears to influence the subcellular targeting and substrate selectivity of AtPAP26. The significant increase in secreted acid phosphatase activity of -Pi seedlings was correlated with the appearance of immunoreactive AtPAP12 and AtPAP26 polypeptides. Analysis of atpap12 and atpap26 T-DNA mutants verified that AtPAP12 and AtPAP26 account for most of the secreted acid phosphatase activity of -Pi wild-type seedlings. Semi-quantitative RT-PCR confirmed that transcriptional controls exert little influence on the up-regulation of AtPAP26 during Pi stress, whereas AtPAP12 transcripts correlate well with relative levels of secreted AtPAP12 polypeptides. We hypothesize that AtPAP12 and AtPAP26 facilitate Pi scavenging from soil-localized organophosphates during nutritional Pi deprivation.

The dual-targeted purple acid phosphatase isozyme AtPAP26 is essential for efficient acclimation of Arabidopsis to nutritional phosphate deprivation.

Induction of intracellular and secreted acid phosphatases (APases) is a widespread response of orthophosphate (Pi)-starved (-Pi) plants. APases catalyze Pi hydrolysis from a broad range of phosphomonoesters at an acidic pH. The largest class of nonspecific plant APases is comprised of the purple APases (PAPs). Although the biochemical properties, subcellular location, and expression of several plant PAPs have been described, their physiological functions have not been fully resolved. Recent biochemical studies indicated that AtPAP26, one of 29 PAPs encoded by the Arabidopsis (Arabidopsis thaliana) genome, is the predominant intracellular APase, as well as a major secreted APase isozyme up-regulated by -Pi Arabidopsis. An atpap26 T-DNA insertion mutant lacking AtPAP26 transcripts and 55-kD immunoreactive AtPAP26 polypeptides exhibited: (1) 9- and 5-fold lower shoot and root APase activity, respectively, which did not change in response to Pi starvation, (2) a 40% decrease in secreted APase activity during Pi deprivation, (3) 35% and 50% reductions in free and total Pi concentration, respectively, as well as 5-fold higher anthocyanin levels in shoots of soil-grown -Pi plants, and (4) impaired shoot and root development when subjected to Pi deficiency. By contrast, no deleterious influence of AtPAP26 loss of function occurred under Pi-replete conditions, or during nitrogen or potassium-limited growth, or oxidative stress. Transient expression of AtPAP26-mCherry in Arabidopsis suspension cells verified that AtPAP26 is targeted to the cell vacuole. Our results confirm that AtPAP26 is a principal contributor to Pi stress-inducible APase activity, and that it plays an important role in the Pi metabolism of -Pi Arabidopsis.

Acid Phosphatase as a Marker for Differentiation of Silkworm (Bombyx Mori L.) Strains

ABSTRACTThis study was carried out on twelve silkworm strains maintained by the Sericulture and Agriculture Experiment Station germplasm bank, located in Vratza, Bulgaria. The polymorphism of acid phosphatase from larval haemolymph was investigated by method of electrophoresis in polyacrylamide gel. Five fundamental types of this enzyme were found in the studied strains of various origins. The acid phosphatase isozymes were considered to be controlled by five codominant alleles. It was found out that the acid phosphatase is very suitable marker for analyzing the inter- and intra-strain diversity and the strain differentiation.

Inheritance of acid phosphatase isozymes in Picea abies

Inheritance of acid phosphatase isozymes in Picea abies

Effect of microbial inoculation during vermicomposting of different organic substrates on microbial status and quantification and documentation of acid phosphatase

In this experiment, three microbial strains were inoculated in two different organic wastes to study their effect on the humic acids content, acid phosphatase activity and microbial properties of the final stabilized products. Pyrophosphate extract of vermicomposts were analyzed through polyacrylamide gel electrophoresis to study the nature of a isozymes in different treatments. Results suggested that vermicomposting increased humic acids content and acid phosphatase activity in organic substrates and microbial inoculation further enhanced the rate of humification and enzyme activity. Although humic acids content in different microorganism-inoculated vermicomposts were statistically at par, acid phosphatase activity in these treatments was significantly (P<0.05) different. Results revealed that microbial respiration was increased due to vermicomposting, but a reduction in microbial biomass was recorded after stabilization of organic wastes. Although vermicomposting increased the value of microbial quotient (qCO(2)), microbial inoculation did not show any significant effect on qCO(2). The zymogram revealed that two isozymes of acid phosphatase (group II and group III) were present in all vermicompost samples and higher acid phosphatase activity in fungi-inoculated vermicomposts might be due to the presence of an additional isozyme (group I) of acid phosphatase.

Differential synthesis of phosphate‐starvation inducible purple acid phosphatase isozymes in tomato (Lycopersicon esculentum) suspension cells and seedlings

This study compares the influence of phosphate (Pi) deprivation on the coordinate synthesis of the principle Pi-starvation inducible (PSI) acid phosphatase (AP) isozymes in a suspension cell culture with the homologous in planta system. Tomato suspension cells express three PSI purple AP isozymes: a heterodimeric intracellular AP (IAP) composed of 63 and 57 kDa subunits, and two monomeric secreted APs (SAPs) (84 kDa SAP1 and 57 kDa SAP2) localized in the culture media. Immunoblots probed with rabbit antibodies raised against purified SAP1 or IAP indicated the immunological distinctiveness of SAP1 relative to IAP and SAP2. Time-course studies of cells and seedlings undergoing a transition from Pi sufficiency to Pi deficiency revealed a close relationship between total IAP or SAP activity and relative amounts of antigenic IAP or SAP polypeptides. Upregulation of the pre-existing IAP in 6-day-old Pi-deficient (-Pi) suspension cells coincided with a 20-fold reduction in intracellular free Pi levels, which occurred 2 d prior to initial accumulation of SAP1 and SAP2 in the culture media. Similarly, root-specific SAP synthesis in -Pi seedlings occurred at least 7 d following IAP induction in roots or shoots. Preferential sequestration of limiting Pi to the leaves of the -Pi seedlings was suggested by the delayed induction of leaf versus root IAP. Our results confirm recent transcript profiling studies suggesting that PSI proteins are subject to both temporal and tissue-specific syntheses in- Pi plants.

Phosphate or phosphite addition promotes the proteolytic turnover of phosphate-starvation inducible tomato purple acid phosphatase isozymes

Within 48 h of the addition of 2.5 mM phosphate (HPO 4 2−, Pi) or phosphite (H 2PO 3 −, Phi) to 8-day-old Pi-starved (−Pi) tomato suspension cells: (i) secreted and intracellular purple acid phosphatase (PAP) activities decreased by about 12- and 6-fold, respectively and (ii) immunoreactive PAP polypeptides either disappeared (secreted PAPs) or were substantially reduced (intracellular PAP). The degradation of both secreted PAP isozymes was correlated with the de novo synthesis of two extracellular serine proteases having M rs of 137 and 121 kDa. In vitro proteolysis of purified secreted tomato PAP isozymes occurred following their 24 h incubation with culture filtrate from Pi-resupplied cells. The results indicate that Pi or Phi addition to −Pi tomato cells induces serine proteases that degrade Pi-starvation inducible extracellular proteins.