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1.
Ph. Looten D. Blanchet J. P. Vandecasteele 《Applied microbiology and biotechnology》1987,25(5):419-425
Summary The -fructofuranosidase activities of a strain of Clostridium acetobutylicum, selected for its capacity to grow on inulinic substrates, were investigated. When grown on inulin, this strain produced extracellular and intracellular -fructofuranosidases, both of which hydrolysed inulin (inulinase activity) and sucrose (invertase activity). Inulinase activity was higher than invertase activity in the extracellular preparation, the opposite being observed for the cellular preparation. The effects of pH and temperature, substrate specificity and the kinetic constants for inulin and sucrose were studied on both preparations, as well as induction by inulin and repression by glucose and fructose of inulinase and invertase activities. The overall results were consistent with the existence of a least one inulinase, (EC 3.2.1.7), mainly but not entirely released in the extracellular medium, and an invertase (3.2.1.26) localized within the cell.Time course hydrolysis experiments of dalhia inulin and Jerusalem artichoke inulofructans by extracellular inulinase showed that this preparation had a remarkably high specificity for hydrolysis of long chain inulofructans. 相似文献
2.
Summary
Bifidobacterium adolescentis ATCC 15703, B. longum ATCC 15707, and B. thermophilum ATCC 25525 were examined for the ability to grow with fructo-oligosaccharides (FOS) as carbohydrate sources. The three species produced cell-associated -fructosidases (inulinases) capable of hydrolysing FOS. Maximum activity was obtained with short-chain FOS with degrees of polymerization (DP) of between three and five (neosugars). The B. thermophilum inulinase was induced by inulin, a long-chain FOS with DP=35, while the enzymes from the other two strains were constitutive. Production of inulinase by all three strains was regulated by catabolite repression. Inulinase activity of the three Bifidobacterium spp. was similar when grown with 0.5% inulin as the carbohydrate source; however, B. thermophilum grew much more rapidly. All three strains utilized crude Jerusalem artichoke flour (JAF) as a carbohydrate source, suggesting that JAF might have commercial application as a food or feed additive to stimulate bifidobacteria in the gut.Contribution no. 802 from the Food Research Centre 相似文献
3.
Irène Efstathiou Gilles Reysset Nicole Truffaut 《Applied microbiology and biotechnology》1986,25(2):143-149
Summary Several strains ofClostridium acetobutylicum, isolated from sugar beet pulps or Jerusalem artichokes, are able to utilize inulin, a -polyfructosane polymer of fructose with glucose as the terminal residue. Inulin-degrading activity, which was detected in cultures of one such strain, ABKn8, grown in Basol-medium containing inulin, reached a maximum at the end of exponential phase. Most of the enzyme activity was detected in the supernatant. It was stably maintained in 0.1 M acetate buffer pH 5.0, and was optimal at pH 4.6. The enzyme, inulinase was induced by inulin, but not by xylose, fructose or sucrose and was repressed by glucose. Inulinase was active against inulin, sucrose and raffinose, but not melezitose. It had a higher affinity for inulin (K
m
: 1.2×10-2 mM) than all the other known inulinases. 相似文献
4.
Total 427 yeast strains from seawater, sediments, mud of salterns, guts of the marine fish, and marine algae were obtained.
After inulinase activity of the yeast cultures was estimated, we found that four strains (OUC1, G7a, OUC2, and G7a1) of the
marine yeasts grown in the medium with inulin could secrete a large amount of inulinase into the medium. The results of routine
identification and molecular methods show that they belong to Pichia guilliermondii OUC1, Cryptococcus aureus G7a, Yarrowia lipolytica OUC2, and Debaryomyces hansenii G7a1, respectively. The optimal pHs of inulinase activity produced by them were 6.0, 5.0, 5.0, and 5.0, respectively, while
the optimal temperatures of inulinase activity produced by them were 60°, 50°, 60°, and 50°C, respectively. A large amount
of monosaccharides and a trace amount of oligosaccharides were detected after the hydrolysis by the crude inulinase produced
by P. guilliermondii OUC1, indicating that the crude inulinase had a high exoinulinase activity while a large amount of monosaccharides and oligosaccharides
were detected after inulin hydrolysis by the crude inulinase produced both by C. aureus G7a and D. hansenii G7a1. However, no monosaccharides and disaccharides were detected after inulin hydrolysis by the crude inulinase produced
by Y. lipolytica OUC2, suggesting that the crude inulinase had no exoinulinase activity. 相似文献
5.
VariousSaccharomyces cerevisiae strains were transformed with a 2 μ-based multicopy expression plasmid, pYIGP, carryingKluyveromyces marxianus inulinase gene under the control ofGAPDH promoter. Among them two strains, SEY2102 and 2805, showed high levels of cell growth and inulinase expression, and were
selected to study their fermentation properties on inulin. Jerusalem artichoke inulin was more effective for cell growth (10∼11
g-dry wt./L at 48 hr) and inulinase expression (1.0 units/mL with SEY2102/pYIGP and 2.5 units/mL with 2805/pYIGP) than other
inulin sources such as dahlia and chicory. It was also found that maximal ethanol production of 9 g/L was obtained from Jerusalem
artichoke inulin at the early stationary phase (around 30 hr), indicating that recombinantS. cerevisiae cells secreting exoinulinase could be used for the simultaneous saccharification of inulin and ethanol fermentation. 相似文献
6.
Expression of exo‐inulinase gene from Aspergillus niger 12 in E. coli strain Rosetta‐gami B (DE3) and its characterization
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Inulin is a linear carbohydrate polymer of fructose subunits (2‐60) with terminal glucose units, produced as carbon storage in selected plants. It cannot directly be taken up by most microorganisms due to its large size, unless prior hydrolysis through inulinase enzymes occurs. The hydrolyzed inulin can be taken up by microbes and/or recovered and used industrially for the production of high fructose syrup, inulo‐oligosaccharides, biofuel, and nutraceuticals. Cell‐free enzymatic hydrolysis would be desirable for industrial applications, hence the recombinant expression, purification and characterization of an Aspergillus niger derived exo‐inulinase was investigated in this study. The eukaroyototic exo‐inulinase of Aspergillus niger 12 has been expressed, for the first time, in an E. coli strain [Rosetta‐gami B (DE3)]. The molecular weight of recombinant exo‐inulinase was estimated to be ~81 kDa. The values of Km and Vmax of the recombinant exo‐inulinase toward inulin were 5.3 ± 1.1 mM and 402.1 ± 53.1 µmol min?1 mg?1 protein, respectively. Towards sucrose the corresponding values were 12.20 ± 1.6 mM and 902.8 ± 40.2 µmol min?1 mg?1 protein towards sucrose. The S/I ratio was 2.24 ± 0.7, which is in the range of native inulinase. The optimum temperature and pH of the recombinant exo‐inulinase towards inulin was 55°C and 5.0, while they were 50°C and 5.5 towards sucrose. The recombinant exo‐inulinase activity towards inulin was enhanced by Cu2+ and reduced by Fe2+, while its activity towards sucrose was enhanced by Co2+ and reduced by Zn2+. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:629–637, 2016 相似文献
7.
Summary One invertase (Inv), five exoinulinases (Exo I; II; III; IV; V) and three endoinulinases (Endo I; II; III) were isolated from a commercial inulinase preparation derived from Aspergillus ficuum using ammonium sulfate precipitation, ion exchange chromatography on DEAE-Sephacel and DEAE-Trisacryl, gel filtration on Ultrogel and Fast Protein Liquid Chromatography on a Mono Q column. The invertase (Inv) had a molecular weight of 84000 and was much more active on sucrose than on inulin: the ratio of activity on inulin and sucrose (I/S ratio) was 0.01. The five exoinulinases show the same molecular weight of 74000 and I/S ratios in the range 0.16–0.36. The three endoinulinases had molecular weight of 64000 and I/S ratios in the range 0.86–2.92. All the -fructofuranosidases were glycoproteins with a high sugar content (from 22 to 41% w/w). A. ficuum is the first described organism containing the three activities: invertase, exo and endoinulinase. 相似文献
8.
Tong Zhang Zhe Chi Zhenming Chi Jean‐Luc Parrou Fang Gong 《Microbial biotechnology》2010,3(5):576-582
It has been confirmed that Saccharomyces sp. W0 can produce high concentration of ethanol. In this study, the INU1 gene cloned from the marine-derived Pichia guilliermondii was transformed into uracil mutant of Saccharomyces sp. W0. The positive transformant Inu-66 obtained could produce 34.2 U ml−1 of extracellular inulinase within 72 h of cultivation. It was found that 15.2 U of inulinase activity per one gram of inulin was suitable for inulin hydrolysis and ethanol production by the transformant Inu-66. During the small-scale fermentation, 13.7 ml of ethanol in 100 ml of medium was produced and 99.1% of the added inulin was utilized by the transformant. During the 2 l fermentation, 14.9% (v/v) of ethanol was produced from inulin and 99.5% of the added inulin was converted into ethanol, CO2 and cell mass. 相似文献
9.
Reconstruction of the spatial structure of inulinase (EC 3.2.1.7) from Kluyveromyces marxianus (an enzyme that hydrolyzes inulin and other fructose-based polymers to fructose) was carried out by highthroughput computational modeling. A structural model of a closely related homologous protein, viz., invertase from yeast Saccharomyces cerevisiae (PDB-ID: 4EQV), was used as a template. The reconstructed model can be used for computer calculations for optimizing the biotechnological feasibility of inulinase. 相似文献
10.
G. Pratap Kumar A. Kunamneni T. Prabhakar P. Ellaiah 《World journal of microbiology & biotechnology》2005,21(8-9):1359-1361
Summary Fifty strains were isolated from different soil samples on synthetic medium containing inulin as a sole carbon source for
the production of extracellular inulinase. Of them, five isolates showed high inulinase activity and one of them was selected
for identification and medium optimization studies. The isolate was identified as Aspergillus niger. Various physical and chemical parameters were optimized for inulinase production. Maximum productivity of inulinase (176 U ml−1) was achieved by employing medium containing 5% (w/v) inulin, galactose as additional carbon source, corn steep liquor and
(NH4)H2PO4 as nitrogen sources, incubation period of 72 h, incubation temperature of 28 °C, pH 6.5, inoculum load at 10% (v/v) level
and medium volume to flask volume ratio of 1:20 (v/v) with indented flasks. 相似文献
11.
Summary A study was made of a β-fructosidase, which is produced extracellularly and intracellularly bySaccharomyces fragilis. The enzyme catalyzes the hydrolysis of inulin, bacterial levans, sucrose, and the fructose portion of raffinose, by splitting
off terminal fructosyl units. It attacks β-2,1 as well as β-2,6 linkages. The enzyme content of inulin-grown cells is sufficient
to allow fermentation of inulin at the same rate as glucose. The ratio of hydrolysis rates with sucrose and inulin was about
25 for the β-fructosidase ofS. fragilis and about 14,000 for invertase.S. fragilis does not contain significant amounts of invertase and it ferments inulin, sucrose and raffinose with the aid of a related,
but different enzyme, inulinase.
Conditions of growth were established which favor inulinase synthesis. Highest yields were obtained with inulin as the carbon
source, and somewhat lower yields with raffinose. Glucose, fructose and sucrose were poor inducers of inulinase. The pH of
the medium during growth on inulin had to be in the range where inulinase could act, otherwise growth was tardy and poor.
In an inulin containing medium aeration favored enzyme production as a result of stimulation of growth. The inulinase content
of the cells in a unit volume was generally greater than that in the culture medium. The intracellular inulinase could be
solubilized quantitatively by autolysis. The intra-and extracellular inulinases were concentrated and purified to the same
extent. Comparison of the two preparations with respect to substrate specificity, rate of inactivation by heat, pH optima
with sucrose (4.2) and with inulin (5.0), and elution patterns from a column of diethylaminoethyl cellulose, indicated that
the intra-and extracellular enzymes were identical. 相似文献
12.
Arun Dev Sharma Prabhjot Kaur Gill Sukhdev Singh Bhullar Prabhjeet Singh 《World journal of microbiology & biotechnology》2005,21(6-7):929-932
Summary An extracellular inulinase (fructanase)-producing strain of Penicillium purpurogenum was isolated from the rhizosphere soil of chicory. Conidia of this selected strain were subjected to simultaneous treatment
with NTG–UV (N-methyl-N′-nitro-N-nitrosoguanidine and ultraviolet radiation) and EtBr–UV (Ethidium bromide–ultraviolet radiation). After mutagenesis, colonies
were screened and among them a few were selected to carry out the inulinase study, which showed a significantly higher inulinase
activity with higher I/S (inulin/sucrose) ratio in all the selected colonies, indicating enhancement of inulinase production
after mutagenic treatments in all the selected mutants. 相似文献
13.
Debaryomyces cantarellii excretes into a buffered medium an inulinase of β-fructofuranosidase type, its synthesis being induced by inulin. The enzyme has a pH optimum at 4 and its optimum temperature is 50°C. ItsK m for inulin is 15mm. 相似文献
14.
Kluyveromyces marxianus NRRL Y-1196 produced the highest inulinase activity (38 U/mg protein) of six yeasts examined after 24 h growth in sauerkraut brine in shaking flasks at 30°C with 0.3% inulin as an enzyme inducer. The enzyme was recovered by acetone fractionation, with a yield of 81%. It had maximum activity at pH 4.4 and 55°C with K
m values for inulin and sucrose of 3.92 mm and 11.9 mm, respectively. The yeast raised the pH from 3.4 to above 7.0, using all the lactic acid in the brine. Growth of K. marxianus in sauerkraut brine with a small amount of inulin may usefully decrease the BOD and concomitantly produce inulinase.The authors are with the Department of Food Science and Technology, Cornell University, Geneva, New York 14456, USA 相似文献
15.
Quang D. Nguyen Frank Mattes Ágoston Hoschke Judit Rezessy-szabó Mahalingeshwara K. Bhat 《Biotechnology letters》1999,21(3):183-186
Aspergillus niger IMI 303386 produced higher levels of intra- and extracellular -fructofuranosidase and inulinase on inulin than on sucrose. Intracellular -fructofuranosidase from sucrose medium catalysed the best transfructosylation reaction. The concentration of fructooligosaccharides (FOS) reached a maximum in 72 h with 25% (w/v) sucrose. The FOS were purified and the main products were kestose and nystose. Inulinase hydrolysed inulin in an exofashion and released mainly fructose. 相似文献
16.
The yeast Kluyveromyces marxianus var. bulgaricus produced large amounts of extracellular inulinase activity when grown on inulin, sucrose, fructose and glucose as carbon
source. This protein has been purified to homogeneity by using successive DEAE-Trisacryl Plus and Superose 6HR 10/30 columns.
The purified enzyme showed a relative molecular weight of 57 kDa by sodium dodecyl sulfate polyacrylamide gel electrophoresis
(SDS-PAGE) and 77 kDa by gel filtration in Superose 6 HR 10/30. Analysis by SDS-PAGE showed a unique polypeptide band with
Coomassie Blue stain and nondenaturing PAGE of the purified enzyme obtained from media with different carbon sources showed
the band, too, when stained for glucose oxidase activity. The optimal hydrolysis temperature for sucrose, raffinose and inulin
was 55°C and the optimal pH for sucrose was 4.75. The apparent K
m values for sucrose, raffinose and inulin are 4.58, 7.41 and 86.9 mg/ml, respectively. Thin layer chromatography showed that
inulinase from K. marxianus var. bulgaricus was capable of hydrolyzing different substrates (sucrose, raffinose and inulin), releasing monosaccharides and oligosaccharides.
The results obtained suggest the hypothesis that enzyme production was constitutive. Journal of Industrial Microbiology & Biotechnology (2000) 25, 63–69.
Received 17 November 1999/ Accepted in revised form 30 May 2000 相似文献
17.
Arthitaya Kawee-ai Nuntinee Ritthibut Apisit Manassa Churairat Moukamnerd Thunnop Laokuldilok Suthat Surawang 《Preparative biochemistry & biotechnology》2018,48(2):194-201
Prebiotic substances are extracted from various plant materials or enzymatic hydrolysis of different substrates. The production of fructo-oligosaccharide (FOS) and inulo-oligosaccharide (IOS) was performed by applying two substrates, sucrose and inulin; oligosaccharide yields were maximized using central composite design to evaluate the parameters influencing oligosaccharide production. Inulin from Jerusalem artichoke (5–15% w/v), sucrose (50–70% w/v), and inulinase from Aspergillus niger (2–7 U/g) were used as variable parameters for optimization. Based on our results, the application of sucrose and inulin as co-substrates for oligosaccharide production through inulinase hydrolysis and synthesis is viable in comparative to a method using a single substrate. Maximum yields (674.82?mg/g substrate) were obtained with 5.95% of inulin, 59.87% of sucrose, and 5.68 U/g of inulinase, with an incubation period of 9?hr. The use of sucrose and inulin as co-substrates in the reaction simultaneously produced FOS and IOS from sucrose and inulin. Total conversion yield was approximately 67%. Our results support the high value-added production of oligosaccharides using Jerusalem artichoke, which is generally used as a substrate in prebiotics and/or bioethanol production. 相似文献
18.
Debaromyces cantarellii Capriotti contains an inulinase activity which is inducible by growth on inulin but not on other β-fructosides. The induction
is inhibited by glucose and fructose. The system is situated in the cell wall and can be best extracted with a 20 mm phosphate
buffer at pH 8.5. The inulinase activity shows pH optima at 4 and 6, suggesting the presence of two enzymes, the latter being
more tightly bound to the cell wall. Both enzymes degrade inulin from the nonreducin end. The cells also contain a constitutive
β-fructofuranosidase with a specificity partly overlapping with that of the inulinase(s). 相似文献
19.
To date, all of microbial inulinases reported showed optimal activity at pH values ranging from 3.5 to 7.0. A bacterial strain,
Marinimicrobium sp. LS-A18, showing high extracellular inulinolytic activity was isolated from a marine solar saltern of the Yellow Sea in
China. Maximum enzyme activity was obtained at 55°C and pH 9.0, respectively. The inulinase activity was induced by inulin,
but not by the other carbon sources employed. Under the optimal medium and culture condition, the highest inulinase activity,
14.6 U/ml, was obtained after 96 h of incubation at shake flask level. The optimal medium for inulinase production was MHI
medium containing 4% inulin, 1% peptone and 5% NaCl, while the optimal culture condition for inulinase production were pH
7.5, temperature 37°C, agitation speed 210 rpm, medium volume 40 ml in 250 ml shake flask, and incubation time 96 h. A large
amount of monosaccharides was released after inulin hydrolysis by the inulinase from strain LS-A18. This is the first report
on alkaline inulinase production from microorganism. 相似文献
20.
The extracellular inulinase structural gene was isolated from the genomic DNA of the marine yeast Pichia guilliermondii strain 1 by PCR. The gene had an open reading frame of 1,542 bp long encoding an inulinase. The coding region of the gene
was not interrupted by any intron. It encoded 514 amino acid residues of a protein with a putative signal peptide of 18 amino
acids and the calculated molecular mass of 58.04 kDa. The protein sequence deduced from the inulinase structural gene contained
the inulinase consensus sequences (WMNXPNGL) and (RDPKVF). It also had ten conserved putative N-glycosylation sites. The inulinase from P. guilliermondii strain 1 was found to be closely related to that from Kluyveromyces marxianus. The inulinase gene without the signal sequence was subcloned into pPICZαA expression vector and expressed in Pichia pastoris X-33. The expressed fusion protein was analyzed by SDS-PAGE and western blotting and a specific band with molecular mass
of about 60 kDa was found. Enzyme activity assay verified the recombinant protein as an inulinase. A maximum activity of 58.7 ± 0.12 U/ml
was obtained from the culture supernatant of P. pastoris X-33 harboring the inulinase gene. A large amount of monosaccharides, disaccharides and oligosaccharides were detected after
the hydrolysis of inulin with the crude recombinant inulinase. 相似文献