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1.
Invertase from a strain of Rhodotorula glutinis 总被引:1,自引:0,他引:1
An invertase (beta-D-fructofuranoside fructohydrolase, EC 3.2.1.26) from Rhodotorula glutinis was purified by ammonium sulfate fractionation, gel filtration and anion exchange chromatography. Invertase molecular weight was estimated to be 100 kDa by analytical gel filtration and 47 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Molecular mass determinations indicated that the native enzyme exists as a homodimer. It is a glycoprotein that contains 19% carbohydrate. The enzyme attacks beta-D-fructofuranoside (raffinose, stachyose and sucrose) from the fructose end. It has a K(m) of 0.227 M and a V(max) of 0.096 micromol/min with sucrose as a substrate. Invertase activity is stable between pH 2.6 and 5.5 for 30 min, maximum activity being observed at pH 4.5. The activation energy was 6520 cal/mol. The enzyme is stable between 20 and 60 degrees C. Mg(2+) and Ca(2+) ions stimulated invertase activity 3-fold, while Fe(2+), K(+), Co(2+), Na(+) and Cu(2+) increased activity about 2-fold. The transfructosylation reaction could not be observed. This enzyme is of particular interest since it appears to have a high hydrolytic activity in 1 M sucrose solution. This fact would make the enzymatic hydrolysis process economically efficient for syrup production using by-products with high salt and sugar contents such as sugar cane molasses. 相似文献
2.
Invertases of Lilium Pollen : Characterization and Activity during In Vitro Germination 总被引:1,自引:1,他引:0
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Two different forms of invertase are found in pollen of lily (Lilium auratum). One form is cytoplasmic (Invertase 1) and the other is bound to the pollen wall (Invertase 2). Invertase 1 has been partially purified and is a glycoprotein (apparent molecular weight, 450 kilodaltons) with a Km of 0.65 millimolar for sucrose. The two invertases differ in pH optimum and thermal stability. Invertases of lily pollen are β-fructofuranosidases which can hydrolyze sucrose but not melizitose. The mature pollen grains have enzyme activity in both cytoplasmic and wall fractions, and no increase in the activity of either occurs during germination. The wall-bound enzyme could not be released by treatments with detergents or high salt concentrations. 相似文献
3.
J.E. Sayago M.A. Vattuone A.R. Sampietro M.I. Isla 《Journal of enzyme inhibition and medicinal chemistry》2013,28(2):123-130
An acid invertase from the fern Pteris deflexa Link was purified and the effect of reaction products on enzyme activity was studied. Fructose and glucose were competitive and non-competitive inhibitors of the enzyme, respectively. Since proteins suppressed glucose and fructose inhibition of the enzyme, an invertase modulation by reaction products is unlikely; nevertheless, an invertase proteinaceous inhibitor previously reported could have a role in this respect. The purified enzyme was an heterodimer M r 90,000 Daltons composed of subunits of 66,000 and 30,000 Daltons. The enzyme had β -fructofuranosidase activity and hydrolyzed mainly sucrose but also raffinose and stachyose, with K m of 3.22, 10.80 and 38.50 mM, respectively. Invertase activity with an optimum pH at 5.0 was present in almost every leaf fern tissue. Pinnas (sporophyll leaflets) had the higher enzyme levels. Invertase histochemical and immunochemical localization studies showed the enzyme mainly in phloem cells. Epidermis, collenchyma and parenchyma cells also showed invertase protein. 相似文献
4.
Sayago JE Vattuone MA Sampietro AR Isla MI 《Journal of enzyme inhibition and medicinal chemistry》2002,17(2):123-130
An acid invertase from the fern Pteris deflexa Link was purified and the effect of reaction products on enzyme activity was studied. Fructose and glucose were competitive and non-competitive inhibitors of the enzyme, respectively. Since proteins suppressed glucose and fructose inhibition of the enzyme, an invertase modulation by reaction products is unlikely; nevertheless, an invertase proteinaceous inhibitor previously reported could have a role in this respect. The purified enzyme was an heterodimer Mr 90,000 Daltons composed of subunits of 66,000 and 30,000 Daltons. The enzyme had beta-fructofuranosidase activity and hydrolyzed mainly sucrose but also raffinose and stachyose, with Km of 3.22, 10.80 and 38.50 mM, respectively. Invertase activity with an optimum pH at 5.0 was present in almost every leaf fern tissue. Pinnas (sporophyll leaflets) had the higher enzyme levels. Invertase histochemical and immunochemical localization studies showed the enzyme mainly in phloem cells. Epidermis, collenchyma and parenchyma cells also showed invertase protein. 相似文献
5.
Comparative Enzymic Studies of Sucrose Metabolism in the Taproots and Fibrous Roots of Beta vulgaris L
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Comparative enzymic studies of sugar beet (Beta vulgaris L.) taproots and fibrous roots revealed differences in invertase (EC 3.2.1.26) and sucrose synthetase (EC 2.4.1.13) activity. Invertase activity of the two root forms differs with respect to specific activity, pH optimum, and enzyme solubility. Acid invertase (pH 4.5) in the taproot was restricted to the peripheral meristematic tissue which produces cells for both taproot and fibrous root growth. This finding supports the hypothesis that the enzyme regulates sucrose partitioning between the taproot and fibrous roots. A distinct alkaline invertase (pH 8.0) was detected in sucrose storage tissues of the taproot. 相似文献
6.
Purification and characterization of three soluble invertases from barley (Hordeum vulgare L.) leaves. 总被引:7,自引:2,他引:5
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Three soluble isoforms of invertase (beta-fructofuranosidase; EC 3.2.1.26) were purified from 7-d-old primary leaves of barley (Hordeum vulgare L.). Invertase I, a monomeric protein of 64 kD, was purified to apparent homogeneity as shown by sodium dodecylsulfate-polyacrylamide gel electrophoresis. Invertases IIA and IIB, multimeric proteins with molecular masses of the 116 and 155 kD, were purified 780- and 1370-fold, respectively, but were not yet homogeneous. Extracts of epidermal strips of leaves contained only invertase IIB. The specific activity of invertase was more than 100-fold higher in the epidermis than in the mesophyll. All three isoforms were acidic invertases, with pH optima of around 5.0 and little activity in the alkaline range. Invertase I had a Km for sucrose of 8.1 mM, and invertases IIA and IIB had much lower values of 1.0 and 1.7 mM, respectively. Invertase I was more than 2-fold more resistant than the other two invertases to the inhibitors HgCl2 and pyridoxal. All three constitutive invertases were found to act also as sucrose-sucrose fructosyltransferases when supplied with high concentrations of sucrose, forming 1-kestose as principal product. However, the fructosyltransferase activity of all three enzymes was inhibited by pyridoxal in the same way as their invertase activity. This characteristic clearly differentiates them from the inducible sucrose-sucrose fructosyltransferase of barley leaves, the activity responsible for the initial steps of fructan biosynthesis, which has previously been shown to be insensitive to pyridoxal. 相似文献
7.
8.
Invertase was entrapped in cellulose triacetate fibers and the properties of the insoluble derivative were studied. Fiber-entrapped invertase was found very stable under operating conditions. For some insoluble preparations a half-life value of 5,300 days was calculated; a sample of invertase fibers, continuously hydrolyzing sucrose, maintained unchanged its activity for five years. The activity displayed by invertase fibers was 15–65% of that of the free enzyme, depending on the amount of entrapped enzyme and on the porosity of the fibers. At very high substrate concentrations the activity of the entrapped invertase approximated to that of the free enzyme. The pH optimum for activity was around 4.5 for the free and entrapped invertase. The native and entrapped enzyme was unstable at temperatures higher than 35°C. The continuous hydrolysis of sucrose using invertase fiber was studied and the potential industrial application of entrapped enzyme is discussed. 相似文献
9.
Invertase covalent grafting onto corn stover 总被引:1,自引:0,他引:1
The covalent coupling of an invertase from baker's yeast onto an agricultural by-product, corn grits, has been developed. The optimal conditions for each step of the chemical modification of the support have been determined: oxidation with sodium metaperiodate, amination with ethylenediamine, reduction with sodium cyanoborohydride, and activation with glutaraldehyde. Activities up to 7.2 x 10(4) mumol reducing sugars produced/min g support could thus be achieved. Invertase coupling onto corn grits yields a derivative with a 25 times higher activity than when coupling this enzyme onto porous silica. The operational stability of invertase immobilized onto corn stover was found to be very high, with a half-life of up to 365 days at 40 degrees C when using a 2M sucrose solution as substrate. This immobilization method could be easily scaled up to the preparation of 10 kg of invertase derivative. 相似文献
10.
George A. Marzluf 《Journal of bacteriology》1973,115(1):146-152
11.
Saccharum officinarum is one of the most cultivated hybrid varieties among the sugarcane varieties. In sugarcane plant sucrose is the
major carbohydrate which can be stored and transported. Different physiological and biochemical studies on this crop report that
invertase activity and sucrose concentration some how are key limiting step in the process of sucrose accumulation. Significant
efforts have been made in relation to the sucrose cycle by altering the sucrose phosphate synthetase, sucrose synthetase and
invertase. In sugarcane two types of invertase enzymes have been reported on the basis of pH and cellular localization. Invertase
breaks the sucrose into hexoses as a source of energy and carbon. It has also been reported that this enzyme is involved in the
process of cell differentiation and plant development. Progress has been made for the understanding of invertase activity and its
role in sugarcane plant. With the help of biotechnology it is possible to target the desired gene with genetic engineering approach
to increase sucrose content by careful manipulation of invertase (enzyme) gene to increase the sucrose yield in sugarcane. Purpose
of this mini review is to high-light the role of invertase in sugarcane and how to overcome sucrose recovery in sugarcane. 相似文献
12.
Graciela L. Salerno 《Physiologia plantarum》1985,64(2):259-264
Sucrose phosphate synthase (UDP-glucose: D-fructose-6-phosphate-2-glucosyl transferase, EC 2.4.1.14), sucrose synthase (UDP-glucose: D-fructose-2-glucosyl transferase, EC 2.4.1.13) and invertase (β-D-fructofuranoside fructohydrolase, EC 3.2.1.26) were measured in toluene permeabilized cells of Chlorella vulgaris Beijerinck. All three activities were detected at all stages of the growth curve; sucrose synthase and sucrose phosphate synthase showed a zone of maximum activity, while invertase increased with time of growth. Sucrose phosphate synthase and sucrose synthase (sucrose synthesis direction) were stimulated by divalent cations and inhibited by UDP. This inhibition could be reversed by Mg2+ or Mn2+ . Sucrose phosphate synthase activity was inhibited by inorganic phosphate and was enhanced by glucose-6-phosphate, but was insensitive to sucrose. Arbutine decreased sucrose synthase activity in both directions. Sucrose cleavage was inhibited by divalent cations and by pyrophosphate. The effects on the enzyme activities of the presence of 2,4-dichlorophenoxyacetic acid (2,4-D), gibberellic acid, abscisic acid and kinetin in the growth medium were investigated. Sucrose synthase activity was practically unaffected by all plant hormones tested, except for the presence of kinetin which stimulated the activity. Sucrose phosphate synthase activity was increased by both kinetin and abscisic acid. The effect of the latter was partially reversed by the presence of gibberellic acid. 2,4-D and kinetin were potent stimulators of invertase activity. 相似文献
13.
AIMS: To characterize the beta-fructofuranosidase of Bifidobacterium infantis ATCC 15697 and to compare it with other bacterial beta-fructofuranosidases. METHODS AND RESULTS: The beta-fructofuranosidase of B. infantis ATCC 15697 was purified 46.8 times over the crude extract by anion exchange chromatography, ultrafiltration and gel filtration. The sequence of 15 amino acid residues of the NH2 terminal was determined. This enzyme was a monomeric protein (Mr 70 kDa) with beta-fructofuranosidase and invertase activities. The isoelectric point was pH 4.3, the optimum pH 6.0 and pKas (4.5 and 7.2) of two active groups were obtained. The activities were inhibited by Hg2+ and p-chloromercuribenzoic acid (pCMB). The optimal temperature was 37 degrees C and activities were unstable at 55 degrees C. beta-fructofuranosidase activity was more efficient than that of invertase with Vm/Km ratios of 0.65 and 0.025 x 10-3 l min(-1) mg(-1), respectively. The enzyme catalyses the hydrolysis of fructo-oligosaccharides, sucrose and inulin at relative velocities of 100, 10 and 6, respectively. CONCLUSIONS: The enzyme of B. infantis ATCC 15697 is an exo-inulinase which has beta-fructofuranosidase and invertase activities. This protein was different from the beta-fructofuranosidase of another strain of B. infantis (B. infantis JCM no. 7007). SIGNIFICANCE AND IMPACT OF THE STUDY: A better knowledge of bacterial beta-fructofuranosidases, especially from bifidobacteria, has been gained. 相似文献
14.
Extracellular invertase from Aspergillus flavus 总被引:1,自引:0,他引:1
An extracellular invertase was induced in cultures of Aspergillus flavus Link during growth in liquid medium that contained sucrose as the sole carbon source. Synthesis of this enzyme was repressed by the addition of glucose or fructose to sucrose-metabolizing cells, and was induced in a glucose or fructose-metabolizing culture by the addition of sucrose. A. flavus invertase had a pH optimum of 6.0 and an apparent Km of approximately 133 mM for sucrose. The enzyme required potassium phosphate for maximum activity, optimum concentration being 250 mM. The enzyme was partially purified by ammonium sulphate precipitation followed by dialysis and separated by molecular exclusion into three components with molecular weights ranging from approximately 40,000 to 55,000. 相似文献
15.
Trypsin (EC 3.4.4.4) was immobilized in low yield on aminoalkylsilylated magnetite (Fe3O4). Better results were obtained when trypsin was immobilized by crosslinking with glutaraldehyde on magnetite. The preparation contained 36 mg protein/g magnetite and the enzyme retained 46% and 11% of esterase and proteolytic activity. Immobilized trypsin was more heat stable than trypsin. Invertase (β-D -fructofuranoside fructohydrolase, EC 3.2.1.26) was cross-linked on magnetite with glutaraldehyde in low yield due to the inactivation of the enzyme. However in the presence of 1% sucrose, the total activity recovered was 79% of the initial activity and the preparation contained 4.4 mg/g of active invertase. Immobilized invertase was less active than invertase when acting on oligosaccharides of the raffinose family. The immobilized enzymes could be easily recovered, from solutions or suspensions, magnetically. 相似文献
16.
Co-ordinated induction of mRNAs for extracellular invertase and a glucose transporter in Chenopodium rubrum by cytokinins 总被引:9,自引:2,他引:9
Extracellular or cell wall invertase is regarded as crucial to supply sink tissues with carbohydrates via an apoplastic pathway. A cell wall invertase from Chenopodium rubrum was purified to homogeneity and the corresponding cDNA encoding CIN1 was identified via peptide sequences. The CIN1 mRNA was found to be highly induced by physiological concentrations of both adenine- and phenylurea-derived cytokinins in suspension culture cells. This was paralleled both by a higher steady-state protein level and a higher enzyme activity of the extracellular invertase. The cytokinin-inducible accumulation of CIN1 mRNA in tissues of C. rubrum plants supports the physiological significance of this regulatory mechanism. In contrast to the extracellular sucrose cleaving enzyme, the mRNA levels of the two putative intracellular invertases CIN2 and CIN3 and of sucrose synthase were not elevated. In addition, it has been found that the accumulation of mRNA for one out of three hexose transporters present in the suspension culture cells is induced co-ordinately with the mRNA for extracellular invertase by cytokinins. It has been shown that this regulatory mechanism results in higher uptake rates both for sucrose, via the hexose monomers, and for glucose. The increased level of both extracellular invertase and hexose transporters and the resulting higher carbohydrate supply are discussed with respect to the control of carbohydrate partitioning by plant hormones and the molecular basis for known physiological cytokinin responses such as the stimulation of cell division. 相似文献
17.
Characterization and distribution of invertase activity in developing maize (Zea mays) kernels 总被引:1,自引:0,他引:1
Invertase ( β -fructofuranoside fructohydrolase, EC 3.2.1.26) activity in developing maize ( Zea mays L. inbred W64A) was separated into soluble and particulate forms. The particulate form was solubilized by treatment with 1 M NaCl or with other salts. However, CaCl2 inhibited invertase activity, and neither detergents nor 0.5 M methyl mannoside were effective in solubilizing the invertase activity. The soluble and particulate invertases were both glycoproteins, both had pH optima of 5.0 and Km values for sucrose of 2.83 and 1.84 m M , respectively. The apparent molecular weight of salt-solubilized invertase was 40 kDa. Gel filtration of the soluble invertase showed multiple peaks with apparent molecular weights ranging from 750 kDa to over 9 000 kDa. Histochemical staining of cell wall preparations for invertase activity suggested that the particulate invertase is associated with the cell wall. Also, nearly all the invertase activity was localized in the basal endosperm and pedicel tissues, which are sites of sugar transport. No invertase activity was found in the upper endosperm, the embryo or in the placento-chalazal tissue. In contrast, sucrose synthase (EC 2.4.1.13) activity was found primarily in the embryo and the upper endosperm, which are areas of active biosynthesis of storage compounds. 相似文献
18.
Invertase (β-D-fructofuranoside fructohydrolase, EC 3.2.1.26) finds major uses in confectionery and in the production of invert syrup. In the present study, we report on invertase production by wild cultures of Saccharomyces cerevisiae. The yeast strains were isolated from dates available in a local market. Five hyperproducing yeast strains (>100- fold higher invertase activity) were kinetically analysed for invertase production. Saccharomyces cerevisiae strain GCA-II was found to be a better invertase-yielding strain than all the other isolates. The values of Qp and Yp/s for GCA-II were economical as compared to other Saccharomyces cultures. The effect of sucrose concentration, rate of invertase synthesis, initial pH of fermentation medium and different organic nitrogen sources on the production of invertase under submerged culture conditions was investigated. Optimum concentrations of sucrose, urea and pH were 3, 0.2 (w/v), and 6 respectively. The increase in the enzyme yield obtained after optimization of the cultural conditions was 47.7%. 相似文献
19.
Carrot (Daucus carota L.) cell suspension cultures grew well when provided with glucose, fructose, sucrose or raffinose. Galactose and melibiose supported less growth unless supplemented with glucose or fructose. In combination with ten different sugar mixtures, 2-deoxy-D-glucose (dGlc) inhibited culture growth. Inhibitory effects of dGlc were more marked with fructose, melibiose, raffinose or mixtures of these sugars in the culture medium. The presence of glucose or galactose reduced the inhibitory effects of dGlc on culture growth. Experiments with radioactive labelled sugars demonstrated that dGLc uptake was greater in the presence of fructose than glucose, and that growth inhibition of dGlc coincided with its uptake. Reduced protein content was also associated with the inhibitory effects of dGlc. Cultured cells contained lower levels of invertase (EC 3.2.1.26) activity during the active phase of culture growth (up to 25 days after subculture) than when growth had peaked and subsequently declined. Acid and alkaline invertase activities were not greatly reduced by exogenous hexoses. Invertase activity was greatest during periods of low protein content in all cultures and was inhibited by dGlc during the latter phases of the culture period. Free intracellular sugars throughout the culture period consisted mainly of glucose and fructose. 相似文献
20.
J.E. Sayago M.A. Vattuone A.R. Sampietro 《Journal of enzyme inhibition and medicinal chemistry》2013,28(6):517-525
Plant invertases play important roles in sucrose metabolism. Cell wall invertase was reported to participate in phloem loading and unloading. Soluble invertases would be involved in hexose level regulation in mature tissues and in stored sucrose utilization within vacuoles. Invertase inhibitory proteins were described as one of the possible mechanisms for invertase activity regulation in some plant species; nevertheless, these proteins were found only in sink tissues, suggesting that this mechanism would not be relevant in the sucrose turnover of leaves. This report describes the purification of invertase from Pteris deflexa fronds and the occurrence of an invertase inhibitory protein in this fern organ, as well as its purification and invertase-inhibitor interactions. The Mr of the invertase and of its inhibitory protein were 90,000 and 18,000, respectively. SDS-PAGE in the presence of 2-mercaptoetanol gave two subunits for the enzyme (Mr=66,000 and 30,000) and only one for the inhibitor. The inhibitor protein is a glycoprotein (12% w/w of neutral sugars) that did not show agglutinating activity like some others, and also showed a high heat stability at pH 5.0. The optimum pH of invertase activity is 5.0, while invertase inhibitory protein caused maximal inhibition at the same pH value. Invertase-inhibitor complex formation occurs in an immediate manner and a protease activity was discarded. The inhibition is non-competitive (Ki=1.5 × 10 ?6 M) without interactions among the binding sites. The complex is slightly dissociable and sucrose was able to partially reduce the inhibitory effect. Up to the present, invertase inhibitory proteins have been found solely in heterotrophic tissues. In this work we demonstrate that this protein is also present in an autotrophic tissue of a lower vascular plant. 相似文献