Acid Phosphatase Isozymes Research Articles

Molecular Cloning of the Type 5, Iron-containing, Tartrate-resistant Acid Phosphatase from Human Placenta

The type 5, iron-containing, tartrate-resistant acid phosphatase (TR-AP) constitutes a relatively minor intracellular isozyme of acid phosphatase in the human that is immunologically related to uteroferrin, a secreted progesterone-induced protein of the porcine uterus. Here, the purification of small amounts of TR-AP from human placenta is described. When a placental lambda gt11 cDNA library was screened with two short 32P-labeled cDNA clones from within the coding region of uteroferrin, a 1412-base pair cDNA was identified that encodes the entire human TR-AP isozyme. This cDNA contains an open reading frame of 969 base pairs, corresponding to a protein of 323 amino acids. A putative signal sequence of 19 amino acids and two potential glycosylation sites are present. The deduced amino acid sequence of the human TR-AP is 85% identical to that of porcine uteroferrin (whose sequence is also reported here in complete form for the first time) and 82% identical to the corresponding regions of a partial amino acid sequence of a bovine spleen phosphoprotein phosphatase. Northern blotting techniques employing a labeled TR-AP cDNA probe revealed the presence of a 1.5-kilobase transcript in white cells from a patient with hairy cell leukemia, in human K562 erythroleukemic cells, and in Epstein-Barr virus-transformed human B-cells, but not in a human T-cell line. Culture of K562 cells in presence of 10(-8) M phorbol 12-myristate 13-acetate ester for 48-72 h enhanced TR-AP activity per cell about 30-fold and led to a corresponding increase in TR-AP mRNA levels.

Characterization of Quince (Cydonia) Cultivars Using Polyacrylamide Gel Electrophoresis

An investigation of available Cydonia (Cydonia oblonga Mill., quince, membrillo) germplasm by isozyme staining of anionic polyacrylamide gradient electrophoresis gels is described. The isozymes of acid phosphatase, esterase, peroxidase and phenol oxidase showed most diversity and usefulness for this purpose. Eleven groups of quince and two groups of x Pyronia (quince-pear crosses) were distinguished by their isozyme patterns. These patterns distinguish between groups of clonal accessions, and the patterns were constant for each accession during the test period (December, 1986 to August, 1987). Thus, these patterns were considered to represent genetic characteristics suitable for identification purposes. The diversity of isozyme patterns was much less than in Corylus and Pyrus populations previously sampled; and less than that of a restricted pool of apple cultivars previously examined.

Promotion of Flowering by DNA Base Analogues and Changes in Acid Phosphatase and Peroxidase Isozyme Composition in Dark-Grown <italic>Arabidopsis thaliana</italic>

Promotion of Flowering by DNA Base Analogues and Changes in Acid Phosphatase and Peroxidase Isozyme Composition in Dark-Grown <italic>Arabidopsis thaliana</italic>

A novel variant of acid phosphatase isozyme from hemolymph of the silkworm, Bombyx mori.

Four electrophoretic variants of acid phosphatase have been already reported in the silkworm hemolymph. We found an electrophoretic variant (type E) which had been designated O (null) type since the distinct band had not been detected. The band E was undiscernible when the substrate (α-Naphthyl phosphate) and chromogen (Fast blue B) were poured simultaneously on the gel after electrophoresis according to the former procedure. However, if the gel was incubated with α-NP then Fast blue B was added, a distinct band E appeared. Quantitative analyses showed that Fast blue B inhibited the isozyme E competitively but other isozymes noncompetitively. Several lines of experimental evidence suggest the isozyme E is not only a charge isomer but also an isozyme showing different kinetic properties with other isozymes. By crossing experiments isozymes A-E were considered to be controlled by codominant alleles, BphA-BphE, respectively.

Immunochemical characterization of human red cell acid phosphatase isozymes.

An immunological study was performed on human red cell acid phosphatase (ACP1) isozymes encoded by different alleles, each of which is expressed as an electrophoretically fast (f) isozyme and a slow (s) isozyme. These isozymes reacted as two immunochemically different groups. Allele-specific reactions were not detected between either the f isozymes or the s isozymes. Quantitation of ACP1 isozymes in red cells by crossed immunoelectrophoresis revealed a phenotype-dependent variation in the concentration of isozyme protein. A simple gene dosage effect was indicated and the ordering of the ACP1 alleles (ACP1*A less than ACP1*B less than ACP1*C less than ACP1*E) was identical to that found for enzyme activity levels. Also, an allele effect on the proportion between s and f isozymes (s/f) was observed; the ordering here was ACP1*B less than ACP1*A less than ACP1*C, which is the same as that reported for the susceptibility to modulation with purines. These variations in isozyme protein levels appear to account for the phenotypic differences in the intensity of the isozyme bands, when activity-stained after electrophoresis, and in the red cell enzyme activity levels. Investigation of two carriers of a Null allele showed no evidence of an aberrant protein product, and half-normal concentrations of enzyme protein were observed in the red cells of these individuals.

Native blot and immunotransfer of human prostatic acid phosphatase isozymes

Agarose gel isoelectrofocusing is used to separate the isozymes of human prostatic acid phosphatase with retention of enzyme activity. The native blotting of the isozymes onto a nitrocellulose membrane increases the sensitivity of the enzyme stain and is suitable for analysis of isozymes in prostate tissue, which contains little nonprostatic acid phosphatase. The specificity of the transfer is increased by treating the membrane with antibody to human prostatic acid phosphatase prior to the transfer. The specificity of the antibody is conferred to the membrane resulting in a transfer specific for prostatic acid phosphatase. The immunotransfer procedure is applicable to serum which contains appreciable amounts of nonprostatic acid phosphatase.

Leishmania donovani: Generation of monospecific antibody reagents to soluble acid phosphatase

Four monoclonal antibodies (McAbs) were generated against the soluble extracellular acid phosphatase (EC 3.1.3.2) (S-AcP) of Leishmania donovani. These were detected in the primary screen using an ELISA with promastigote culture supernatants as antigen. Three of the McAbs demonstrated bound S-AcP from such culture supernatants in an enzyme activity binding assay. All immunoprecipitated metabolically labeled S-AcP but none showed any binding to the promastigote surface by indirect immunofluorescence. Moreover, none reacted with Triton X-100 solubilized plasma membranes by immunoprecipitation or Western blotting. These results demonstrated that the McAbs did not recognize the surface membrane bound acid phosphatase, but were specific for the extracellular soluble enzyme. Further, none of the antibodies immunoprecipitated any of the five human acid phosphatase isozymes or reacted with them in Western blots or the enzyme activity binding assay. Therefore, they are specific for the parasite-derived enzyme. One of these was used to affinity purify sufficient L. donovani S-AcP to immunize a rabbit and generate a specific, polyvalent antiserum. This polyvalent antibody immunoprecipitated S-AcP activity but did not cross-react with the surface membrane acid phosphatase, indicating that these two parasite enzymes are separate gene products.

Enzyme activity and phytohormone production of a mycorrhizal fungus, Laccarialaccata

Six isolates of Laccarialaccata, S-167 from a forest nursery and S-238, S-283, S-326, S-444, and S-472 from natural forests, were analyzed for acid phosphatase, alkaline phosphatase, and nitrate reductase activity, acid phosphatase isozyme patterns, and IAA and cytokinin production. Differences in enzyme activity and phytohormone production were prominent among the isolates. The patterns of acid phosphatase isozyme could be clearly divided into three host-related groups. Two polymorphic gene loci could be identified as coding for enzymes of acid phosphatase. Two of these gene loci, Acp-b and Acp-c, are characterized by mostly constant, host-dependent frequencies. The other, Acp-d, exhibits allele frequencies related to different habitats. Five isolates share the same Acp-a habitat. Four isolates share the Acp-b habitat, two the Acp-c habitat, and only one isolate, S-238, was from a high elevation at the Acp-d habitat. The isolate from a forest nursery differed strikingly in several characteristics from the other isolates, all of which were from a natural forest. This suggests that nursery soilmanagement practices may select for particular edaphic ecotypes of mycorrhizal fungi.

Enzymes and Growth Substances of Rhizopogon Species in Relation to Mycorrhizal Hosts and Infrageneric Taxonomy

Acid and alkaline phosphatase and nitrate reductase activities, acid phosphatase isozyme patterns, and cytokinins and indoleacetic acid production were determined for thirteen isolates encompassing six species and three sections of the genus Rhizopogon. Two of the species are mycorrhizal associates particularly with Douglas-fir, one with pines, one a host generalist but particularly associated with pines, and two broadly host-general. Isolates within species generally differed as much from each other as did species or sections of the genus. Of all parameters examined, isozyme patterns were most consistent within species and differed most between sections.

Enzymes and Growth Substances of Rhizopogon Species in Relation to Mycorrhizal Hosts and Infrageneric Taxonomy

Acid and alkaline phosphatase and nitrate reductase activities, acid phosphatase isozyme patterns, and cytokinins and indoleacetic acid production were determined for thirteen isolates encompassing six species and three sections of the genus Rhizopogon. Two of the species are mycorrhizal associates particularly with Douglas-fir, one with pines, one a host generalist but particularly associated with pines, and two broadly host-general. Isolates within species generally differed as much from each other as did species or sections of the genus. Of all parameters examined, isozyme patterns were most consistent within species and differed most between sections.

HIV antigen in the brains of patients with the AIDS dementia complex.

Human immunodeficiency virus infection was identified immunohistochemically in the brains of 8 patients with acquired immune deficiency syndrome dementia complex. Using a monoclonal antibody against a structural viral protein (p25), infection was detected in white matter and basal ganglia in a distribution paralleling that of the major neuropathological abnormalities. Viral antigen was identified principally in perivascular and parenchymal macrophages and in multinucleated cells of macrophage origin that were identified morphologically and by immunocytochemical staining for acid phosphatase isozyme. In 4 of the 8 patients, viral antigen was also detected in acid-phosphatase-negative, process-bearing neuroglial cells; in 2 patients, antigen was detected in basal ganglion cells that were morphologically consistent with neurons and in alkaline-phosphatase-positive cells with elongated nuclei that were most likely of endothelial origin.

Activities and Isozymes of Acid Phosphatase in Schizophyllum commune: A Re-Examination

Activities and Isozymes of Acid Phosphatase in Schizophyllum commune: A Re-Examination

Activities and Isozymes of Acid Phosphatase inSchizophyllum Commune: A Re-Examination

Specific activities and isozymes of acid phosphatase (AcPase) were analyzed in colonies of Schizophyllum commune, a basidiomycete. Isogenic strains examined included two homokaryons, the dikaryon formed from them, and Bmut, a homokaryon with a primary mutation in the B incompatibility factor. In the homokaryons and the dikaryon AcPase activity increased with age to a peak at nine days and then declined; however, the activity of Bmut continued to rise after the ninth day. Two bands of activity were always present in polyacrylamide gels of extracts from all four strains at all ages analyzed. However, a third band appeared at different times in aging colonies of the homokaryons versus the dikaryon. A plot of the log of the mobility of the isozymes versus gel concentration provided evidence that these isozymes were primarily charge isomers, hence, not likely the result of proteolysis. These results are in contrast to the numerous isozymes of AcPase observed by Wang and Raper for several strains of Schizophyllum. We think their studies contain artifacts produced by the procedures they used.

Characterization of Filbert (Corylus) Species and Cultivars Using Gradient Polyacrylamide Gel Electrophoresis

A chemical identification procedure previously used to identify apple and pear species, cultivars and clonal accessions, was tried with Corylus (filbert, hazel) species, cultivars and clonal accessions. Following electrophoresis, the peroxidase, phenol oxidase, and acid phosphatase isozyme patterns on anionic polyacrylamide gradient gels were determined. These patterns were found to vary between clonal accessions, but did not change, within a given accession during and following the test period (May through October). Thus, these patterns were considered to represent genetic characteristics suitable for identification purposes. The patterns were used to identify 78 Corylus accessions at the National Clonal Germplasm Repository Corvallis, Oregon. All accessions tested (species, cultivars and clones) were distinguishable using this system. The diversity of isozyme patterns was greater in Corylus than Pyrus populations previously sampled. This technique appears to have the potential to readily identify filbert accessions and could be an important aid in the characterization of germplasm material.

Leaf acid phosphatase isozymes in the diagnosis of phosphorus status in field-grown wheat

This study examined the inorganic and total phosphorus in wheat plant tops, along with the phosphatase activity and the phosphatase isozymes occurring in the youngest fully expanded leaves, from two field experiments done in 1983 and I984 at two sites in South Australia. Twelve levels of phosphorus were used in the first year, and split plots in the second year allowed the effects of current and/or residual phosphorus treatments to be determined.The inorganic or total phosphorus concentration, or content per plant, could be related to plant and grain yield, but values relative to phosphorus deficiency varied between different stages of growth and between sites. Relationships with inorganic phosphorus were no better than those with total phosphorus.Leaf acid phosphatase assays were a better measure, providing similar values relative to plant deficiency between sites, between seasons and irrespective of whether current or residual phosphorus was the source, but the values varied with plant age.The phosphatase zymograms identified an isozyme which appeared when plants were deficient in phosphorus. It did not appear when they were adequately supplied. The isozyme appeared at all stages of growth to anthesis, and from late tillering through to heading was related to grain yield at both sites, in both seasons and whether freshly applied or residual phosphorus was the source of supply. For diagnostic purposes, specific sets of standards would be required for the phosphorus and phosphatase assays. They would not be required for the phosphatase zymograms.

Comparison of eight Pisolithustinctorius isolates for growth rate, enzyme activity, and phytohormone production

Eight isolates of Pisolithustinctorius (Pers.) Coker and Couch (three each from Georgia and northern California and one each from Oregon and Washington) were compared invitro for growth rate, for alkaline and acid phosphatase and nitrate reductase activities, for acid phosphatase isozyme patterns, and for cytokinin, indoleacetic acid, and gibberellin production. Significant differences appeared between isolates for each parameter examined. All isolates showed relatively low phosphatase and nitrate reductase activities. Isolate S-359 from northern California grew the slowest in culture and produced significantly more indoleacetic acid than all other isolates and more cytokinin than six of the other seven isolates; this isolate was also the only one of the eight that did not share at least one acid phosphatase allele with the others in the isozyme analysis.

Biochemical studies on initiation of callus in Solanum surattense

Summary Activities of peroxidase, acid phosphatase and α-amylase and total phenols increased just before appearance of callus. Starch and reducing sugars accumulated soon after inoculation of explants to the callus forming medium and decreased subsequently. Five new isoperoxidases appeared during induction period out of which two disappeared before initiation of callus. One new isozyme of acid phosphatase also appeared prior to the appearance of callus.

Identification of Apple Rootstock Cultivars by Isozyme Analysis

Abstract Thirty-three clonal apple (Malus domestica Borkh.) rootstocks were characterized by their peroxidase, esterase, acid phosphatase, and idoleacetic acid oxidase isozyme banding patterns. Polyacrylamide disk-gel electrophoresis of shoot bark protein extracts, with a 5% to 15% acrylamide concentration gradient in the separating gel, provided adequate resolution for the anionic isozymes for all 4 enzymes, while a 5% to 20% gradient was used to separate the cationic isozymes of peroxidase and acid phosphatase. Growing conditions (greenhouse and 3 different field locations) and sample timing (February, May, August, and November) did not affect banding patterns. Virus-tested clones showed isozymic differences compared with the original contaminated material. Tissue culture propagated selections from EMLA 7 and MAC 9 stoolbeds expressed different banding patterns compared to original stocks.

Fingerprinting Apple Cultivars by Electrophoretic Isozyme Banding Patterns

Anionic electrophoretic isozyme patterns of peroxidase, esterase and acid phosphatase and cationic peroxidase isozyme patterns from shoot bark protein extracts were used to identify clonal apple scion cultivars. Each of the 21 cultivars included in this study developed a unique combination of isozyme patterns which allowed it to be distinguished from the others. Sports within cultivars exhibited identical patterns of enzymes, with the exception of ‘Wijcik’, a natural compact mutant of ‘McIntosh’ which could be distinguished from the latter, although it was indistinguishable from the cultivar ‘Spartan’. Isozyme patterns remained constant when samples were taken from wood of different ages, at several times of the year and with trees growing in different locations and on different rootstocks.

Characterization of Pyrus Species and Cultivars Using Gradient Polyacrylamide Gel Electrophoresis

A chemical identification procedure previously used to identify apple clones was tried with pear species and clones. Following electrophoresis, the peroxidase, esterase, and acid phosphatase isozyme patterns on anionic polyacrylamide gradient gels were determined. These patterns were found to vary with the species and clone, but not to change, within a clone during the growing season. Thus, these patterns were considered to represent genetic characteristics. The patterns were used to identify 37 selected Pyrus accessions at the National Clonal Germplasm Repository, Corvallis, Oregon. All species tested were distinguishable using this system. All the accessions of P. calleryana selected from the NCGR collection were distinct: however, one clone from outside the collection had an identical pattern to one inside the collection. Among the Chinese pear clones (complex hybrids of P. ussuriensis × P. pyrifolia) tested, three pairs of clones had the same combination. This technique appears to have the potential to readily identify pear specimens, and could be an important aid in the characterization of germplasm material.