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1.
Cellulomonas flavigena strain KU (ATCC 53703) is a cellulolytic, Gram-positive bacterium which produces large quantities of an insoluble exopolysaccharide (EPS) when grown in minimal media with a high carbon-to-nitrogen (C/N) ratio. Earlier studies proved the EPS is structurally identical to the linear β-1,3-glucan known as curdlan and provided evidence that the EPS functions as a carbon and energy reserve compound. We now report that C. flavigena KU also accumulates two intracellular, glucose-storage carbohydrates under conditions of carbon and energy excess. These carbohydrates were partially purified and identified as the disaccharide trehalose and a glycogen/amylopectin-type polysaccharide. A novel method is described for the sequential fractionation and quantitative determination of all three carbohydrates from culture samples. This fractionation protocol was used to examine the effects of C/N ratio and osmolarity on the accumulation of cellular carbohydrates in batch culture. Increasing the C/N of the growth medium caused a significant accumulation of curdlan and glycogen but had a relatively minor effect on accumulation of trehalose. In contrast, trehalose levels increased in response to increasing osmolarity, while curdlan levels declined and glycogen levels were generally unaffected. During starvation for an exogenous source of carbon and energy, only curdlan and glycogen showed substantial degradation within the first 24 h. These results support the conclusion that extracellular curdlan and intracellular glycogen can both serve as short-term reserve compounds for C. flavigena KU and that trehalose appears to accumulate as a compatible solute in response to osmotic stress. 相似文献
2.
De Baets S Du Laing S François C Vandamme EJ 《Journal of industrial microbiology & biotechnology》2002,29(4):181-184
In liquid culture conditions, the yeast-like fungus Tremella mesenterica occurs in the yeast state and synthesizes an exopolysaccharide (EPS) capsule, which is eventually released into the culture
fluid. It is composed of an α-1,3-D-mannan backbone, to which β-1,2 side chains are attached, consisting of D-xylose and D-glucuronic acid. Potato dextrose broth (PDB) seemed to be an excellent medium for both growth of the yeast cells and synthesis
of the EPS. This medium is composed solely of an extract of potatoes to which glucose was added. Yet an important disadvantage
of this production medium is the presence of starch in the potato extract, since Tremella cells are not capable of metabolizing this component; furthermore, it coprecipitates upon isolation of the polymer [3]. In
this respect, it was essential to remove the starch in order to achieve high polysaccharide production and recovery. A good
method was the removal of starch through ultrafiltration of the PDB medium before inoculation of the strain. This resulted
in an excellent starch-free medium in which other components essential for polysaccharide production were still present [3].
Through implementation of single and cyclic fed-batch fermentations with glucose feed, 1.6- and 2.2-fold increases in EPS
yield were obtained, respectively. Lowering the carbon source level by using a cyclic fed-batch technique might decrease the
osmotic effect of glucose or any catabolite regulation possibly exerted by this sugar on enzymes involved in EPS synthesis.
Journal of Industrial Microbiology & Biotechnology (2002) 29, 181–184 doi:10.1038/sj.jim.7000276
Received 18 March 2002/ Accepted in revised form 20 May 2002 相似文献
3.
The genus Cellulomonas is comprised of a group of Gram-positive, soil bacteria capable of utilizing cellulose as their sole source of carbon and energy. Cellulomonas flavigena KU was originally isolated from leaf litter and subsequently shown to produce large quantities of a curdlan-type (-1,3-glucan) exopolysaccharide (EPS) when provided with an excess of glucose or other soluble carbon-source. We report here that curdlan EPS is also produced by Cellulomonas flavigena KU when growing on microcrystalline cellulose in mineral salts-yeast extract media. Microscopic examination of such cultures shows an adherent biofilm matrix composed of cells, curdlan EPS, and numerous surface structures resembling cellulosome complexes. Those Cellulomonas species that produce curdlan EPS are all non-motile and adhere to cellulose as it is broken down into soluble sugars. These observations suggest two very different approaches towards the complex process of cellulose degradation within the genus Cellulomonas. 相似文献
4.
Sushma Deepthi Arli U. B. Trivedi K. C. Patel 《World journal of microbiology & biotechnology》2011,27(6):1415-1422
Cellulomonas flavigena UNP3, a natural isolate from vegetable oil contaminated soil sample has been studied for growth associated exopolysaccharide
(EPS) production during growth on glucose, groundnut oil and naphthalene. The EPS showed matrix formation surrounding the
cells during scanning electron microscopy. Cell surface hydrophobicity and emulsifying activity studies confirmed the role
of EPS as bioemulsifier. Emulsifying activity was found to increase with time (0.2 U/mg for 10 min to 0.27 U/mg for 30 min).
Emulsification index, E24 value increased with the increase in EPS concentration. Degradation of polyaromatic hydrocarbons was confirmed using gas
chromatography analysis. FTIR analysis showed presence of characteristic absorbance at 895.10 cm−1 for β-configuration of glucan. NMR studies also revealed EPS produced by C. flavigena UNP3 as a linear β-1, 3-d-glucan, and a curdlan like polysaccharide. 相似文献
5.
Exopolysaccharides (EPS) from lactic acid bacteria contribute to specific rheology and texture of fermented milk products
and finds applications even in non-dairy foods and in therapeutics. Box-Behnken model of response surface methodology (RSM)
was employed to formulate the production medium for exopolysaccharide (EPS). FT-IR spectral analysis of the purified EPS from
Lactobacillus plantarum MTCC 9510 revealed prominent characteristic groups corresponding to polyhydric alcohols. The degradation temperature (Td)
of the polysaccharide was found to be 260°C with the help of thermo gravimetric analysis (TGA). Structure elucidation of the
EPS showed that it consists of a trisaccharide repeating unit of α-d-glucose, β-d-glucose and α-d-mannose. 相似文献
6.
The aerobic nitrogen fixing xylanolytic bacterium Paenibacillus pabuli strain ATSKP produces loosely attached capsular polysaccharide KP-EPS. On 0.5% birchwood xylan 70 ± 5.02 mg of KP-EPS was
produced per gram dry weight of cells by the fourth day of growth in the absence of combined nitrogen source at 30°C. It was
separated and purified using centrifugation, cold acetone precipitation and dialysis and is a sulfate containing heteropolymer
as revealed by FT-IR spectrometry and elemental analysis. CHN analysis revealed the presence of 37.50% carbon, 5.90% hydrogen
and 8.28% nitrogen in KP-EPS. Absence of phosphorus was confirmed by 31P NMR. ICP-OES analysis showed the presence of various metals in small concentrations. Specific binding with aniline blue
suggested the presence of (1,3)-β-d-glucan. Thermal gravimetric analysis and differential scanning calorimetric analysis confirmed its thermal stability as high
as 200°C. The EPS was not pseudo plastic and the viscosity was less than xanthan. The intrinsic viscosity did not reduce drastically
when dissolved in 0.1 M NaCl. 相似文献
7.
A C S Rizzatti J A Jorge H F Terenzi C G V Rechia M L T M Polizeli 《Journal of industrial microbiology & biotechnology》2001,26(3):156-160
A β-D-xylosidase was purified from cultures of a thermotolerant strain of Aspergillus phoenicis grown on xylan at 45°C. The enzyme was purified to homogeneity by chromatography on DEAE-cellulose and Sephadex G-100. The
purified enzyme was a monomer of molecular mass 132 kDa by gel filtration and SDS-PAGE. Treatment with endoglycosidase H resulted
in a protein with a molecular mass of 104 kDa. The enzyme was a glycoprotein with 43.5% carbohydrate content and exhibited
a pI of 3.7. Optima of temperature and pH were 75°C and 4.0–4.5, respectively. The activity was stable at 60°C and had a K
m of 2.36 mM for p-nitrophenyl-β-D-xylopiranoside. The enzyme did not exhibit xylanase, cellulase, galactosidase or arabinosidase activities. The purified enzyme
was active against natural substrates, such as xylobiose and xylotriose. Journal of Industrial Microbiology & Biotechnology (2001) 26, 156–160.
Received 23 June 2000/ Accepted in revised form 29 September 2000 相似文献
8.
M-K Kim I-Y Lee J-H Lee K-T Kim Y-H Rhee Y-H Park 《Journal of industrial microbiology & biotechnology》2000,25(4):180-183
We investigated the influence of inorganic phosphate concentration on the production of curdlan by Agrobacterium species. A two-step culture method was employed where cells were first cultured, followed by curdlan production under nitrogen-limiting
conditions. In the curdlan production step, cells did not grow but metabolized sugar into curdlan. Shake-flask experiments
showed that the optimal phosphate concentration for curdlan production was in the range of 0.1–0.3 g l−1. As the cell concentration increased from 0.42 to 1.68 g l−1 in shake-flask cultures, curdlan production increased from 0.44 to 2.80 g l−1. However, the optimal phosphate concentration range was not dependent upon cell concentration. The specific production rate
was about 70 mg curdlan g-cell−1 h−1 irrespective of cell concentration. When the phosphate concentration was maintained at 0.5 g l−1 under nitrogen-limiting conditions, as high as 65 g l−1 of curdlan was obtained in 120 h. Journal of Industrial Microbiology & Biotechnology (2000) 25, 180–183.
Received 25 October 1999/ Accepted in revised form 21 July 2000 相似文献
9.
Vargas-García MC López MJ Elorrieta MA Suárez F Moreno J 《Journal of industrial microbiology & biotechnology》2002,29(3):129-133
The relationship between exopolysaccharide (EPS) production by Azotobacter vinelandii ATCC 12837 from 4-hydroxybenzoic acid as sole carbon source and other physiological parameters was investigated. In relation
to growth, Azotobacter needed more time in 4-hydroxybenzoic acid to reach levels of biomass similar to those obtained when sugars were used, although
the phenolic compound led to a more extensive exponential phase. The encystment process was initiated after cells had grown
for 24 h, in which small amounts of EPS were synthesized and poly-β-hydroxybutyrate (PHB) accumulation began. Both polymers,
EPS and PHB, showed a similar evolution with time, as well as the formation of cysts, which points out the existence of a
relation between these parameters. This was corroborated by a statistical study, in which significant correlations (P<0.05) were observed when each parameter was compared to the two others. Journal of Industrial Microbiology & Biotechnology (2002) 29, 129–133 doi:10.1038/sj.jim.7000288
Received 01 February 2002/ Accepted in revised form 13 June 2002 相似文献
10.
B C Saha 《Journal of industrial microbiology & biotechnology》2001,27(4):241-245
An extracellular β-xylosidase from a newly isolated Fusarium verticillioides (NRRL 26518) was purified to homogeneity from the culture supernatant by concentration by ultrafiltration using a 10,000
cut-off membrane, ammonium sulfate precipitation, DEAE Bio-Gel A agarose column chromatography and SP-Sephadex C-50 column
chromatography. The purified β-xylosidase (specific activity, 57 U/mg protein) had a molecular weight (mol. wt.) of 94,500
and an isoelectric point at pH 7.8. The optimum temperature and pH for action of the enzyme were 65°C and 4.5, respectively.
It hydrolyzes xylobiose and higher xylooligosaccharides but is inactive against xylan. The purified β-xylosidase had a K
m value of 0.85 mM (p-nitrophenol-β-D-xyloside, pH 4.5, 50°C) and was competitively inhibited by xylose with a K
i value of 6 mM. It did not require any metal ion for activity and stability. Journal of Industrial Microbiology & Biotechnology (2001) 27, 241–245.
Received 20 May 2001/ Accepted in revised form 06 July 2001 相似文献
11.
Deep-sea hydrothermal vents: A new source of innovative bacterial exopolysaccharides of biotechnological interest? 总被引:2,自引:0,他引:2
Guezennec J 《Journal of industrial microbiology & biotechnology》2002,29(4):204-208
Polysaccharides and, in particular, microbial polysaccharides represent a class of important products of growing interest
for many sectors of industry. Although many known marine bacteria produce exopolysaccharides (EPS), continuation in looking
for new polysaccharide-producing microorganisms is promising. Hydrothermal deep-sea vents could be a source of novel EPS as
indicated by the screening of a number of mesophilic heterotrophic bacteria recovered from different locations. Although originating
from such extreme environment, some bacteria were shown to biosynthesize innovative EPS under laboratory conditions. Their
specific rheological properties either in the presence or absence of monovalent and divalent ions, biological activities,
metal binding capabilities, and novel chemical composition mean that these EPS are expected to find many applications in the
near future. Journal of Industrial Microbiology & Biotechnology (2002) 29, 204–208 doi:10.1038/sj.jim.7000298
Received 18 March 2002/ Accepted in revised form 13 June 2002 相似文献
12.
For the production of α-D-glucose-1-phosphate (G-1-P), α-1,4-D-glucan phosphorylase from Thermus caldophilus GK24 was partially purified to a specific activity of 13 U mg−1 and an enzyme recovery of 15%. The amount of G-1-P reached maximum (18%) when soluble starch was used as substrate, and the
smallest substrate for G-1-P formation was maltotriose. The structure of purified G-1-P was confirmed by comparison to 13C-NMR data for an authentic sample. In addition to G-1-P, glucose-6-phosphate (12%) was simultaneously produced when 10 mM
maltoheptaose was used as substrate. Journal of Industrial Microbiology & Biotechnology (2000) 24, 89–93.
Received 12 May 1999/ Accepted in revised form 29 August 1999 相似文献
13.
Exopolysaccharide (EPS) production was compared among three strains of lactobacilli. Lactobacillus rhamnosus strain 9595M can be classified among the highest EPS-producing strains of lactic acid bacteria reported to date with a maximum
EPS production of 1275 mg L−1. Under controlled pH, no significant differences in the quantity of EPS produced could be detected between carbon source
(glucose or lactose) or fermentation temperature (32 or 37°C). In milk, strains ATCC 9595M and R produced more than 280 mg L−1 EPS whereas strain Type V produced less than 80 mg L−1 EPS. Journal of Industrial Microbiology & Biotechnology (2000) 24, 251–255.
Received 10 September 1999/ Accepted in revised form 22 December 1999 相似文献
14.
Hayashi S Ohno T Ito M Yokoi H 《Journal of industrial microbiology & biotechnology》2001,26(5):276-279
β-Xylosidase was extracted from Aureobasidium sp. ATCC 20524 and purified to homogeneity. The molecular mass was estimated at 411 kDa. The enzyme contained 15.3% (w/w)
carbohydrate. The optimum pH and temperature were pH 3.5 and 80°C, respectively. The enzyme was stable at pH 3.5–9 after 3
h and at 80°C after 15 min. The Michaelis constant (K
m) and maximum velocity (V
max) toward p-nitrophenyl-β-D-xyloside were 2.0 mmol l−1 and 0.94 mmol min−1 mg−1 protein, respectively. The enzyme was inhibited strongly by mercury, lead, and copper ions. Journal of Industrial Microbiology & Biotechnology (2001) 26, 276–279.
Received 02 August 2000/ Accepted in revised form 15 December 2000 相似文献
15.
Biosynthesis of the (1,3)-β-d-glucan (curdlan) in Agrobacterium sp., is believed to proceed by the repetitive addition of glucosyl residues from UDP-glucose by a membrane-embedded curdlan
synthase (CrdS) [UDP-glucose: (1,3)-β-d-glucan 3-β-d-glucosyltransferase; EC 2.4.1.34]. The catalytic module of CrdS (cm-CrdS) was expressed in good yield from a cDNA encoding
cm-CrdS cloned into the pET-32a(+) vector, containing a coding region for thioredoxin, and from the Champion™ pET SUMO system
that possesses a coding region of a small ubiquitin-related modifier (SUMO) partner protein. The two DNA fusions, designated
pET-32a_cm-CrdS and SUMO_cm-CrdS were expressed as chimeric proteins. High yields of inclusion bodies were produced in E. coli and these could be refolded to form soluble proteins, using a range of buffers and non-detergent sulfobetaines. A purification
protocol was developed, which afforded a one-step on-column refolding and simultaneous purification of the recombinant 6xHis-tagged
SUMO_cm-CrdS protein. The latter protein was digested by a specific protease to yield intact cm-CrdS in high yields. The refolded
SUMO_cm-CrdS protein did not exhibit curdlan synthase activity, but showed a circular dischroism spectrum, which had an α/β-type-like
conformation. Amino acid sequences of tryptic fragments of the SUMO_cm-CrdS fusion and free cm-CrdS proteins, determined by
MALDI/TOF confirmed that the full-length proteins were synthesized by E. coli, and that no alterations in amino acid sequences occurred. A three-dimensional model of cm-CrdS predicted the juxtaposition
of highly conserved aspartates D156, D208, D210 and D304, and the QRTRW motif, which are likely to play roles in donor and
acceptor substrate binding and catalysis. 相似文献
16.
von Weymarn N Kiviharju K Leisola M 《Journal of industrial microbiology & biotechnology》2002,29(1):44-49
Ten heterofermentative lactic acid bacteria were compared in their ability to produce D-mannitol from D-fructose in a resting state. The best strain, Leuconostoc mesenteroides ATCC-9135, was examined in high cell density membrane cell-recycle cultures. High volumetric mannitol productivity (26.2
g l−1 h−1) and mannitol yield (97 mol%) were achieved. Using the same initial biomass, a stable high-level production of mannitol was
maintained for 14 successive bioconversion batches. Applying response surface methodology, the temperature and pH were studied
with respect to specific mannitol productivity and yield. Moreover, increasing the initial fructose concentration from 100
to 120 and 140 g l−1 resulted in decreased productivities due to both substrate and end-product inhibition of the key enzyme, mannitol dehydrogenase
(MDH). Nitrogen gas flushing of the bioconversion media was unnecessary, since it did not change the essential process parameters.
Journal of Industrial Microbiology & Biotechnology (2002) 29, 44–49 doi:10.1038/sj.jim.7000262
Received 12 November 2001/ Accepted in revised form 30 March 2002 相似文献
17.
An extracellular endo-d-arabinase enzyme produced by the bacterial strain of Cellulomonas was purified 77.1-fold with 0.20% recovery for protein by DEAE Sepharose anion exchange, Sephacryl S-300 gel filtration and
blue Sepharose affinity chromatography, and designated as CEDAase. The apparent molecular mass of CEDAase was 45 kDa determined
by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. CEDAase is an endoenzyme for arabinogalactan with the main and
specific product of hexa-arabinofuranoside. It reacts optimally with its substrate, arabinogalactan, at approximately pH 8.0
and at 40 °C. CEDAase shows stability in the pH range of 6.0–9.0 and at the temperature below 50 °C. The Km measured for the CEDAase was 55.6 μM, with an apparent Vmax of 0.083 μmol/min. To our knowledge, for the first time, the current work obtains an extracellular Cellulomonas endo-d-arabinase enzyme that might be potentially served as a tool enzyme for hydrolyzing specific cell wall such as Mycobacterium cell. It is purified as an important potential initial material basis for mass spectrometric sequencing and chemical gene
synthesis. It may make it possible to clone and express this valuable endo-d-arabinase and make it available to the mycobacteria scientific community. 相似文献
18.
S Chakraborti R K Sani U C Banerjee R C Sobti 《Journal of industrial microbiology & biotechnology》2000,24(1):58-63
An extracellular β-galactosidase which catalyzed the production of galacto-oligosaccharide from lactose was harvested from
the late stationary-phase of Bacillus sp MTCC 3088. The enzyme was purified 36.2-fold by ZnCl2 precipitation, ion exchange, hydrophobic interaction and gel filtration chromatography with an overall recovery of 12.7%.
The molecular mass of the purified enzyme was estimated to be about 484 kDa by gel filtration on a Sephadex G-200 packed column
and the molecular masses of the subunits were estimated to be 115, 86.5, 72.5, 45.7 and 41.2 kDa by sodium dodecyl sulfate-polyacrylamide
gel electrophoresis. The isoelectric point of the native enzyme, determined by polyacrylamide gel electrofocusing, was 6.2.
The optimum pH and temperature were 8 and 60°C, respectively. The Michaelis–Menten constants determined with respect to o-NO2-phenyl-β-D-galactopyranoside and lactose were 6.34 and 6.18 mM, respectively. The enzyme activity was strongly inhibited (68%) by galactose,
the end product of lactose hydrolysis reaction. The β-galactosidase was specific for β-D anomeric linkages. Enzyme activity was significantly inhibited by metal ions (Hg2+, Cu2+ and Ag+) in the 1–2.5 mM range. Mg2+ was a good activator. Catalytic activity was not affected by the chelating agent EDTA. Journal of Industrial Microbiology & Biotechnology (2000) 24, 58–63.
Received 09 February 1999/ Accepted in revised form 24 September 1999 相似文献
19.
Elin Säwén Eine Huttunen Xue Zhang Zhennai Yang Göran Widmalm 《Journal of biomolecular NMR》2010,47(2):125-134
The use of lactic acid bacteria in fermentation of milk results in favorable physical and rheological properties due to in
situ exopolysaccharide (EPS) production. The EPS from S. thermophilus ST1 produces highly viscous aqueous solutions and its structure has been investigated by NMR spectroscopy. Notably, all aspects
of the elucidation of its primary structure including component analysis and absolute configuration of the constituent monosaccharides
were carried out by NMR spectroscopy. An array of techniques was utilized including, inter alia, PANSY and NOESY-HSQC TILT
experiments. The EPS is composed of hexasaccharide repeating units with the following structure: → 3)[α-d-Glcp-(1 → 4)]-β-d-Galp-(1 → 4)-β-d-Glcp-(1 → 4)[β-d-Galf-(1 → 6)]-β-d-Glcp-(1 → 6)-β-d-Glcp-(1 →, in which the residues in square brackets are terminal groups substituting backbone sugar residues that consequently
are branch-points in the repeating unit of the polymer. Thus, the EPS consists of a backbone of four sugar residues with two
terminal sugar residues making up two side-chains of the repeating unit. The molecular mass of the polymer was determined
using translational diffusion experiments which resulted in Mw = 62 kDa, corresponding to 64 repeating units in the EPS. 相似文献
20.
I-Y Lee W T Seo G J Kim M K Kim C S Park Y H Park 《Journal of industrial microbiology & biotechnology》1997,18(4):255-259
Maltose and sucrose were efficient carbon sources for the production of curdlan by a strain of Agrobacterium sp. A two-step, fed-batch operation was designed in which biomass was first produced, followed by curdlan production which
was stimulated by nitrogen limitation. There exists an optimal timing for nitrogen limitation for curdlan production in the
two-step, fed-batch operation. Maximum curdlan production (60 g L−1) was obtained from sucrose with a productivity of 0.2 g L−1 h−1 when nitrogen was limited at a cell concentration of 16.0 g L−1. It was also noted that the curdlan yield from sucrose was as high as 0.45 g curdlan g−1 sucrose, and the highest specific production rate was 1.0 g curdlan g−1 cells h−1 right after nitrogen limitation. Of particular importance was the use of molasses as a cheap carbon source to produce curdlan
in the two-step, fed-batch cultivation. As high as 42 g L−1 of curdlan with a yield of 0.35 g curdlan g−1 total sugar was obtained after 120 h of fed-batch cultivation.
Received 20 August 1996/ Accepted in revised form 26 November 1996 相似文献