Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.2.1.15 (pectinase)
 2,440 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

1. A mild, reproducible extraction procedure, using 0.5% ammonium oxalate, was developed for the isolation of polysaccharides containing d-apiose from the cell wall of Lemna minor. On a dry-weight basis the polysaccharide fractions extracted with ammonium oxalate made up 14% of the material designated cell walls and contained 20% of the d-apiose originally present in the cell walls. The cell walls, as isolated, contained 83% of the d-apiose present in L. minor. 2. After extraction with ammonium oxalate, purified polysaccharides were obtained by DEAE-Sephadex column chromatography and by fractional precipitation with sodium chloride. With these procedures the material extracted at 22 degrees C could be separated into at least five polysaccharides. On a dry-weight basis two of these polysaccharides made up more than 50% of the material extracted at 22 degrees C. There was a direct relationship between the d-apiose content of the polysaccharides and their solubility in sodium chloride solutions; those of highest d-apiose content were most soluble. 3. All the polysaccharides isolated appeared to be of one general type, namely galacturonans to which were attached side chains containing d-apiose. The d-apiose content of the apiogalacturonans varied from 7.9 to 38.1%. The content of esterified d-galacturonic acid residues in all apiogalacturonans was low, being in the range 1.0-3.5%. Hydrolysis of a representative apiogalacturonan with dilute acid resulted in the complete removal of the d-apiose with little or no degradation of the galacturonan portion. 4. Treatment of polysaccharide fractions with pectinase established that those of high d-apiose content and soluble in m-sodium chloride were not degraded, whereas those of low d-apiose content and insoluble in m-sodium chloride were extensively degraded. When the d-apiose was removed from a typical pectinase-resistant polysaccharide, the remainder of the polysaccharide was readily degraded by this enzyme. 5. Periodate oxidation of representative polysaccharide fractions and apiogalacturonans and determination of the formaldehyde released showed that about 50% of the d-apiose molecules were substituted at either the 3- or the 3'-position.
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PMID:Isolation and partial characterization of apiogalacturonans from the cell wall of Lemna minor. 431 31

Alcohol insoluble solids from apple were extracted in sequence by buffer at 20 degrees C and at 70 degrees C, EDTA/oxalate, and mild alkali, yielding four populations of pectins. These pectins and the insoluble residue were characterized by their sugar composition, degree of esterification (methyl ester and O-acetyl groups), molecular weight distribution, and degradability by the combination of endopolygalacturonase (PG) and pectin esterase (PE) and by rhamnogalacturonase (RGase) after chemical saponification. After PG/PE treatment, the remaining high molecular weight material representing the pectic hairy regions was isolated and characterized. Clear differences were found in the sugar composition of the fractions obtained, while only small variations were observed in the sugar linkage composition. The pectic hairy regions were further degraded by RGase and the digests separated into high molecular weight and oligomeric degradation products. These "RGase oligomers" consisted of between 4 and 9 sugar units with a backbone of alternating rhamnose and galacturonic acid residues, partly substituted with galactose linked to C-4 of the rhamnose moiety. Both the absolute amount of RGase oligosaccharides released as well as the degree of galactose-substitution of the oligomers increased when severer extraction conditions were used. Relatively more RGase oligomers were released from the low molecular weight hairy regions as compared to the high molecular weight fraction. Typical high molecular weight fragments isolated from the RGase digests of various hairy regions included residual segments of the rhamnogalacturonan backbone rich in arabinose and a polymer presumably enriched in xylogalacturonan segments.
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PMID:Different populations of pectic hairy regions occur in apple cell walls. 852 28

A protocol for extracting polysaccharides from cell walls has been modified and used to analyze histochemically two fruits with opposite characteristics. Grapes are nonclimacteric fruits and are harvested at full maturity. In contrast, kiwi fruits are climacteric and are harvested and consumed before they are physiologically mature. The two fruits were analyzed histochemically using two protocols. One method is defined as chemical, and is based on subsequential extractions of pectins by chemical agents. The other is defined as enzymatic because it removes pectins using pectinase followed by hot ammonium oxalate. In both protocols, two types of hemicellulosic polymers are removed by 1 M and 4 M/KOH leaving a cellulosic residue on the slide. Both protocols remove the same amount of pectins, thus confirming their precision. The sum of hemicellulose and the cellulosic insoluble residue are equivalent using the two methods, but the relative amounts of the cellulose and hemicellulosic polymers were dependent upon the method of extraction. When the enzyme was used to extract the pectins, there was less cellulose and more hemicellulose. The removal of polysaccharides by ammonium oxalate and by guanidinethiocyanate in the enzymatic and the chemical protocols, respectively, yielded approximately the same amount of removed material. Similar results were obtained from both fruits. Grape, being softer than kiwi fruit, was relatively richer in pectic substances and less rich in hemicellulose and cellulose polymers. No difference in cell wall material could be ascribed to the different ripening habits.
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PMID:Comparative histochemical analysis of cell wall polysaccharides by enzymatic and chemical extractions of two fruits. 906 6

The transfer of a methyl group from S-adenosyl-L-methionine onto the carboxyl group of alpha-1,4-linked-galactosyluronic acid residues in the pectic polysaccharide homogalacturonan (HGA) is catalyzed by an enzyme commonly referred to as pectin methyltransferase. A pectin methyltransferase from microsomal membranes of tobacco (Nicotiana tabacum) was previously characterized (F. Goubet, L.N. Council, D. Mohnen [1998] Plant Physiol 116: 337-347) and named HGA methyltransferase (HGA-MT). We report the solubilization of HGA-MT from tobacco membranes. Approximately 22% of the HGA-MT activity in total membranes was solubilized by 0.65% (w/v) 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonic acid containing 1 mM dithioerythritol. The addition of phosphatidylcholine and the methyl acceptors HGA or pectin (30% degree of esterification) to solubilized enzyme increased HGA-MT activity to 35% of total membrane-bound HGA-MT activity. Solubilized HGA-MT has a pH optimum of 7.8, an apparent K(m) for S-adenosyl-L-methionine of 18 microM, and an apparent V(max) of 0. 121 pkat mg(-1) of protein. The apparent K(m) for HGA and for pectin is 0.1 to 0.2 mg mL(-1). Methylated product was solubilized with boiling water and ammonium oxalate, two conditions used to solubilize pectin from the cell wall. The release of 75% to 90% of the radioactivity from the product pellet by mild base treatment showed that the methyl group was incorporated as a methyl ester rather than a methyl ether. The fragmentation of at least 55% to 70% of the radiolabeled product by endopolygalacturonase, and the loss of radioactivity from the product by treatment with pectin methylesterase, demonstrated that the bulk of the methylated product produced by the solubilized enzyme was pectin.
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PMID:Solubilization and partial characterization of homogalacturonan-methyltransferase from microsomal membranes of suspension-cultured tobacco cells. 1048 84

Water-soluble polysaccharide fractions VO1-VO4 were isolated from the squeezed berries of snowball tree (Viburnum opulus) by successive extraction with water at various temperatures and pH and with aqueous solutions of ammonium oxalate. These fractions were purified by ion-exchange chromatography on DEAE cellulose, and the homogeneity of the purified polysaccharides was determined by gel filtration on Sephacryl S-500. Acidic polysaccharides close to pectins in their sugar composition were found in all the extracts (fractions VO1-1, VO2-1, VO3-2, and VO4-2). Residues of galacturonic acid, galactose, arabinose, and (to a lesser extent) rhamnose are their main constituents. Neutral polysaccharides composed mainly of galactose and mannose residues were additionally found in fractions extracted with acidified water (pH 4.0) and with aqueous ammonium oxalate solutions. Partial acidic hydrolysis and digestion with pectinase of acidic polysaccharides indicated that their carbohydrate backbone consists of alpha-1,4-linked residues of D-galacturonic acid. NMR spectra of acidic polysaccharides (fractions VO3-2 and VO3-3) confirmed this and demonstrated that their side oligosaccharide chains are composed of beta-1,4-linked galactopyranose residues and of terminal and 2,5- and 3,5-substituted residues of alpha-arabinofuranose at a Gal: Ara ratio of 3:1. Some polysaccharides from V. opulus were found to possess an immunostimulating activity: they enhance phagocytosis, in particular, the phagocytic index and the secretion of lysosomal enzymes with peritoneal macrophages. Calcium ions were found to be necessary for the appearance of the stimulating effect of acidic polysaccharides from V. opulus.
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PMID:[The isolation, preliminary study of structure and physiological activity of water-soluble polysaccharides from squeezed berries of Snowball tree Viburnum opulus]. 1080 53

Using extraction with 0.75% aqueous ammonium oxalate, the following polysaccharide fractions were isolated: tanacetans TVF, TVS, and TVR from floscules, sprouts, and roots, respectively, of Tanacetum vulgare L., spread throughout the European North of Russia. The sugar chain of tanacetan TVF consists of D-galacturonic acid (61.4%), arabinose (14.7%), galactose (10.2%), and rhamnose (3.7%) as the main constituents as well as xylose, glucose, mannose, apiose, and 2-O-methylxylose in trace amounts. Tanacetans TVS and TVR were shown to differ in the sugar quantitative composition. They contain 67 and 28% galacturonic acid, respectively. A partial acid hydrolysis of the tanacetan TVF gave a polysaccharide fragment TVF1, alpha-1,4-D-galacturonan (GalA 98.2%). Digestion with pectinase (alpha-1,4-D-polygalacturonase) resulted in fragment TVF3, containing residues of arabinose (27.1%) and galactose (17.3%). NMR spectroscopy allowed detection of the terminal residues of alpha-Araf and beta-Galp as well as of the residues of alpha-Araf substituted in 3,5- and 5-positions. Thus, tanacetan TVF was proved to be a pectic polysaccharide.
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PMID:[Isolation and characterization of polysaccharides from Tansy]. 1125 43

The synergistic activities of oxalic acid and endopolygalacturonases are thought to be essential for full virulence of Sclerotinia sclerotiorum and other oxalate-producing plant pathogens. Both oxalic acid production and endopolygalacturonase activity are regulated by ambient pH. Since many gene products with pH-sensitive activities are regulated by the PacC transcription factor in Aspergillus nidulans, we functionally characterized a pacC gene homolog, pac1, from S. sclerotiorum. Mutants with loss-of-function alleles of the pac1 locus were created by targeted gene replacement. In vitro mycelial growth of these pac1 mutants was normal at acidic pH, but growth was inhibited as culture medium pH was increased. Development and maturation of sclerotia in culture was also aberrant in these pac1 replacement mutants. Although oxalic acid production remained alkaline pH-responsive, the kinetics and magnitude of oxalate accumulation were dramatically altered. Additionally, maximal accumulation of endopolygalacturonase gene transcripts (pg1) was shifted to higher ambient pH. Virulence in loss-of-function pac1 mutants was dramatically reduced in infection assays with tomato and Arabidopsis. Based on these results, pac1 appears to be necessary for the appropriate regulation of physiological processes important for pathogenesis and development of S. sclerotiorum.
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PMID:The Sclerotinia sclerotiorum pac1 gene is required for sclerotial development and virulence. 1297 2

The necrotrophic fungal pathogen Sclerotinia sclerotiorum secretes oxalic acid and endo-polygalacturonase (endo-PG) in host plants. Oxalic acid acidifies the plant tissue to values more suitable to endo-PG activity. However, we observed that the infected soybean seedlings possessed a pH of 3.8, which is below that optimal for endo-PG activity (4.5 to 5.0). We investigated, therefore, the effects of pH (from 5.0 to 3.6) and oxalate (5 to 20 mM) on the activity of the major basic endo-PG (PGb) and towards an acidic endo-PG (PGa) secreted by S. sclerotiorum during soybean infection. We verified that only PGb activity is stimulated by oxalate, while at the lowest pH levels, PGa escapes the inhibition of a soybean polygalacturonase-inhibiting protein (PGIP). These results, performed on polygalacturonic acid, were apparently consistent with data obtained from studies with soybean hypocotyl segments, in which PGb activity was increased by oxalate and PGa maintained its activity also at pH 3.6, possibly because at this pH the PGIP contained in the plant tissue is inactive. Reverse transcription-polymerase chain reaction analysis showed that, during soybean infection, the expression of the putative pga gene is delayed in comparison to the basic one. The different temporal expressions of the two endo-PGs and their differing responses to pH, oxalate, and PGIP seem to be consistent with a possible maximization of the fungal PG activity in the host tissue.
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PMID:Relationships among endo-polygalacturonase, oxalate, pH, and plant polygalacturonase-inhibiting protein (PGIP) in the interaction between Sclerotinia sclerotiorum and soybean. 1559 46

Botrytis cinerea is a necrotrophic pathogen that attacks more than 200 plant species. Here, the nonpathogenic mutant A336, obtained via insertional mutagenesis, was characterized. Mutant A336 was nonpathogenic on leaves and fruits, on intact and wounded tissue, while still able to penetrate the host plant. It grew normally in vitro on rich media but its conidiation pattern was altered. The mutant did not produce oxalic acid and exhibited a modified regulation of the production of some secreted proteins (acid protease 1 and endopolygalacturonase 1). Culture filtrates of the mutant triggered an important oxidative burst in grapevine (Vitis vinifera) suspension cells, and the mutant-plant interaction resulted in the formation of hypersensitive response-like necrosis. Genetic segregation analyses revealed that the pathogenicity phenotype was linked to a single locus, but showed that the mutated gene was not tagged by the plasmid pAN7-1. Mutant A336 is the first oxalate-deficient mutant to be described in B. cinerea and it differs from all the nonpathogenic B. cinerea mutants described to date.
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PMID:Characterization of a new, nonpathogenic mutant of Botrytis cinerea with impaired plant colonization capacity. 1662 75

A pectin polysaccharide named bergenan was isolated from the freshly collected leaves of the leather bergenia Bergenia crassifolia by extraction with an aqueous solution of ammonium oxalate. The main component of its carbohydrate chain was shown to be the residues of D-galacturonic acid (about 80%). In addition, the polysaccharide contains residues of galactose, arabinose, and rhamnose; their total content is less than 15%. It was shown that the bergenan samples from bergenia leaves collected at different vegetation periods (from July to September) do not substantially differ either in monosaccharide composition or in the viscosity of aqueous solutions they form. The results of enzymatic hydrolysis by alpha-1,4-galacturonase (pectinase), partial acidic hydrolysis, NMR spectroscopy, and methylation with subsequent analysis of the results by GC-MS indicate that the bergenan macromolecule contains the regions of a linear alpha--1,4-D-galactopyranosyluronan and rhamnogalacturonan-I (RG-1). Galacturonan responds for a greater part of the macromolecule. A considerable amount of its constituent galacturonic acid residues are present as methyl esters. The side chains in RG-I are attached to the rhamnopyranose residues of the main carbohydrate chain by 1,4-link and are composed of the residues of terminal arabinofuranose and galactopyranose, 1,5-linked (-arabinofuranose, and 1,4-and 1,6-linked beta-galactopyranose. The branching points of the side chains of the RG-I molecule are 3,4- and 3,6-di-O-substituted galactose residues.
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PMID:[Structural study of bergenan, a pectin from Bergenia crassifolia]. 1737 59


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