共查询到20条相似文献,搜索用时 15 毫秒
1.
H. Sztajer W. Wang H. Lünsdorf A. Stocker R. D. Schmid 《Applied microbiology and biotechnology》1996,45(5):600-606
Geotrichum candidum was found to produce a lactate oxidase. The enzyme was purified by gel filtration and ion-exchange chromatography. The purified
lactate oxidase showed a molecular mass of 50 kDa under denaturing and about 400 kDa under non-denaturing conditions. Transmission
electron micro-scopy analysis confirmed an octameric structure. FMN was found to be a cofactor for this enzyme. Polarographic
studies confirmed an oxygen uptake by the lactate oxidase. The enzyme showed specificity towards the L isomer of lactate and
did not oxidise pyruvate, fumarate, succinate, maleate and ascorbate. It was stable at alkaline pH and also for 15 min at
45°C. The addition of glycerol and dextran 500 000 to the enzyme sample enhanced storage stability.
Received: 28 September 1995/Received revision: 10 January 1996/Accepted: 15 January 1996 相似文献
2.
In our previous studies, the yeast Endomyces fibuliger LU677 was found to degrade amygdalin in bitter apricot seeds. The present investigation shows that E. fibuliger LU677 produces extracellular β-glycosidase activity when grown in malt extract broth (MEB). Growth was very good at 25 °C
and 30 °C and slightly less at 35 °C. When grown in MEB of pH 5 and pH 6 with addition of 0, 10 or 100 ppm amygdalin, E. fibuliger produced only slightly more biomass at pH 5, and was only slightly inhibited in the presence of amygdalin. Approximately,
60% of the added amygdalin was degraded (fastest at 35 °C) during an incubation period of 5 days. Supernatants of cultures
grown at 25 °C and pH 6 for 5 days were tested for the effects of pH and temperature on activity (using amygdalin, linamarin
and prunasin as substrates). Prunase activity had two pH optima (pH 4 and pH 6), amygdalase and linamarase only one each at
pH 6 and pH 4–5 respectively. The linamarase activity evolved earlier than amygdalase (2 days and 4 days respectively). The
data thus indicate the presence of at least two different glycosidases having different pH optima and kinetics of excretion.
In the presence of amygdalin, lower glycosidase activities were generally produced. However, the amygdalin was degraded from
the start of the growth, strongly indicating an uptake of amygdalin by the cells. The temperature optimum for all activities
was at 40 °C. Activities of amygdalase (assayed at pH 4) and linamarase (at pH 6) evolving during the growth of E. fibuliger were generally higher in cultures grown at 25 °C and 30 °C. TLC analysis of amygdalin degradation products show a two-stage
sequential mechanism as follows: (1) amygdalin to prunasin and (2) prunasin to cyanohydrin.
Received: 16 September 1997 / Received revision: 6 October 1997 / Accepted: 14 October 1997 相似文献
3.
Y Takashima Y Yamaga S Mitsuda 《Journal of industrial microbiology & biotechnology》1998,20(3-4):220-226
A novel thermophilic Bacillus smithii strain SC-J05-1, isolated from a hot spring, had the ability of hydrating nitrile to form amide. The nitrile hydratase was
purified to homogeneity from the microbial cells of SC-J05-1 and was characterized. The enzyme was a 130-kDa protein composed
of two different subunits (25.3 kDa and 26.8 kDa) and contained cobalt ions. This enzyme had the optimal temperature of 40°C
and was stable up to 50°C. The optimal pH was in the alkaline region higher than pH 10.
Received 02 September 1997/ Accepted in revised form 06 February 1998 相似文献
4.
T. Kobayashi Y. Hakamada J. Hitomi K. Koike S. Ito 《Applied microbiology and biotechnology》1996,45(1-2):63-71
Alkalophilic Bacillus sp. KSM-K16 produced three alkaline proteases, as detected by polyacrylamide gel electrophoresis (PAGE). The major protease,
designated M protease, was recently purified to homogeneity and its properties were characterized. In the present study, two
minor proteases, designated H protease and N protease, were purified to homogeneity from cultures of this organism. H protease
had a molecular mass of 28 kDa, as estimated by sodium dodecyl sulfate/PAGE (SDS-PAGE) and its maximum activity against casein
was observed at pH 11.0 and at 55°C. N protease consisted of two polypeptide chains with molecular masses of 12.5 kDa and 14.5 kDa, as estimated by SDS-PAGE,
although it migrated as a single protein band during non-denaturing PAGE. Its maximum activity was observed at pH 11.0 and
at 60°C. The amino-terminal sequences of H protease and of the 14.5-kDa polypeptide of N protease were identical to that of M protease.
The electrophoretic relationship between the three enzymes was examined after they had been stored at different pH values
and at 5°C. M protease was converted to H protease more rapidly at pH 11 than at pH 8 or below, and H protease was converted to M protease
at pH 8 or below but not at pH 11. N protease appeared to be the autolytic product of the M and H proteases.
Received: 12 December 1994/Received last revision: 9 June 1995/Accepted: 31 July 1995 相似文献
5.
M. Graf A. Brunella M. Kittelmann K. Laumen O. Ghisalba 《Applied microbiology and biotechnology》1997,47(6):650-657
A new amidohydrolase deacetylating several N-acetyl-1-phenylethylamine derivatives (R)-specifically was found in Arthrobacter aurescens AcR5b. The strain was isolated from a wet haystack by enrichment culture with (R)-N-acetyl-1-phenylethylamine as the sole carbon source. (R) and (S )-N-acetyl-1-phenylethylamine do not serve as inducers for acylase formation. By improving the growth conditions the enzyme production
was increased 47-fold. The amidohydrolase was purified to homogeneity leading to a 5.2-fold increase of the specific activity
with a recovery of 67%. A molecular mass of 220 kDa was estimated by gel filtration. Sodium dodecyl sulfate/polyacrylamide
gel electrophorosis shows two subunits with molecular masses of 16 kDa and 89 kDa. The optimum pH and temperature were pH 8
and 50 °C, respectively. The enzyme was stable in the range of pH 7–9 and at temperatures up to 30 °C. The enzyme activity
was inhibited by Cu2+, Co2+, Ni2+, and Zn2+, and this inhibition was reversed by EDTA.M
Received: 20 September 1996 / Received version: 23 December 1996 / Accepted: 30 December 1996 相似文献
6.
β-Fructofuranosidase was purified from commercial alkaline protease (Aspergillus oryzae origin). The optimal pH of its transfructosylating activity was more alkaline (pH 8) than that of its hydrolyzing activity
(pH 5). In the case of a 24-h reaction with sucrose, the hydrolysis and transfructosylation reaction were optimal at pH 4–5
and pH 8, respectively. In the reaction at pH 8 1-kestose and nystose were the main fructooligosaccharides produced. The transfer
ratio was hardly different between pH 5 and pH 8 early in the reaction, but the transfer products (1-kestose and nystose)
were decreased at pH 5 as the reaction proceeded because of their hydrolysis.
Received: 18 January 1995/Received last revision: 23 August 1995/Accepted: 13 September 1995 相似文献
7.
Bacillus subtilis CBTK 106, isolated from banana wastes, produced high titres of α-amylase when banana fruit stalk was used as substrate in
a solid-state fermentation system. The effects of initial moisture content, particle size, cooking time and temperature, pH,
incubation temperature, additional nutrients, inoculum size and incubation period on the production of α-amylase were characterised.
A maximum yield of 5 345 000 U mg-1 min-1 was recorded when pretreated banana fruit stalk (autoclaved at 121 °C for 60 min) was used as substrate with 70% initial
moisture content, 400 μm particle size, an initial pH of 7.0, a temperature of 35 °C, and additional nutrients (ammonium sulphate/sodium
nitrate at 1.0%, beef extract/peptone at 0.5%, glucose/sucrose/starch/maltose at 0.1% and potassium chloride/sodium chloride
at 1.0%) in the medium, with an inoculum-to-substrate ratio of 10% (v/w) for 24 h. The enzyme yield was 2.65-fold higher with
banana fruit stalk medium compared to wheat bran.
Received: 18 April 1995/Received last revison: 6 May 1996/Accepted: 9 May 1996 相似文献
8.
A number of nutritional factors influencing growth and glucose oxidase (EC 1.1.3.4) production by a newly isolated strain
of Penicillium pinophilum were investigated. The most important factors for glucose oxidase production were the use of sucrose as the carbon source,
and growth of the fungus at non-optimal pH 6.5. The enzyme was purified to apparent homogeneity with a yield of 74%, including
an efficient extraction step of the mycelium mass at pH 3.0, cation-exchange chromatography and gel filtration. The relative
molecular mass (M
r) of native glucose oxidase was determined to be 154 700 ± 4970, and 77 700 for the denatured subunit. Electron-microscopic
examinations revealed a sandwich-shaped dimeric molecule with subunit dimensions of 5.0 × 8.0 nm. Glucose oxidase is a glycoprotein
that contains tightly bound FAD with an estimated stoichiometry of 1.76 mol/mol enzyme. The enzyme is specific for d-glucose, for which a K
m value of 6.2 mM was determined. The pH optimum was determined in the range pH 4.0–6.0. Glucose oxidase showed high stability
on storage in sodium citrate (pH 5.0) and in potassium phosphate (pH 6.0), each 100 mM. The half-life of the activity was
considerably more than 305 days at 4 °C and 30 °C, and 213 days at 40 °C. The enzyme was unstable at temperatures above 40 °C
in the range pH 2.0–4.0 and at a pH above 7.0.
Received: 18 November 1996 / Received revision: 3 March 1997 / Accepted: 7 March 1997 相似文献
9.
G. C. Passador-Gurgel S. A. Furlan J. K. Melier R. Jonas 《Applied microbiology and biotechnology》1996,45(1-2):158-161
Fourteen strains of yeast from genera Kluyveromyces, Candida, Debaryomyces and Schizosaccharomyces were investigated for inulinase production. In the first stage, the microtitre reader system SLT was used for the determination
of enzyme activity and the evaluation of cellular growth. Different culture conditions were tested and four strains of Kluyveromyces were selected on the basis of enzyme activity and growth capacity at low pH and high temperature: K. marxianus CBS 6397, DSM 70792, ATCC 36907 and IZ 619. These strains were tested in greater volume using pH 4.0, 45°C and inulin (10 g/l) as selection conditions. On the basis of results obtained, the strain K. marxianus ATCC 36907 was selected for inulinase production. Enzyme stability at low pH (4.0) as well as high temperature (50°C) for 10, 30 and 60 min was also evaluated, but no significant difference in enzyme activity was observed. It could be demonstrated
that the microtitre reader system is an excellent method for the screening of microorganisms.
Received: 31 May 1995/Received revision: 20 September 1995/Accepted: 29 September 1995 相似文献
10.
L. Li T. A. Freier P. A. Hartman J. W. Young D. C. Beitz 《Applied microbiology and biotechnology》1995,43(5):887-892
A resting-cell assay was established to evaluate the cholesterol reductase activity of Eubacterium coprostanoligenes ATCC 51222. Cell suspensions from cholesterol-free media rapidly reduced cholesterol to coprostanol. Optimal assay conditions
in a 1-ml reaction mixture were determined to be up to 1 h of incubation and up to 0.25 mg bacterial protein/assay with at
least 1 mM cholesterol as substrate. The cholesterol reductase activity in cells decreased as a function of storage time at
22°C, 4°C and −20°C. Filling the headspace of the reaction mixture with H2 increased the activity about 20%. Optimal cholesterol reductase activity occurred at pH 7.5 in sodium phosphate buffer. Pyruvate
and reducing agents in the buffer increased the activity. This study has validated assay conditions for determination of cholesterol
reductase activity in resting cells of E. coprostanoligenes.
Received: 2 August 1994/Received revision: 15 November 1994/Accepted: 8 December 1994 相似文献
11.
V. Zverlov K. Piotukh O. Dakhova G. Velikodvorskaya R. Borriss 《Applied microbiology and biotechnology》1996,45(1-2):245-247
The nucleotide sequence of the xynA gene, encoding extracellular xylanase A of Thermotoga neapolitana, was determined. The xynA gene was 3264 base pairs (bp) long and encoded a putative polypeptide of 1055 amino acids. Three different domains were identified
by sequence comparison and functional analysis of proteins with N- and/or C-terminal deletions. The core domain displayed
significant homology to members of the glycosyl hydrolase family 10. N- and C-terminal domains were dispensable for enzymatic
activity and seemed to be responsible for thermostability and cellulose binding, respectively. The intact gene and its truncated
variants were expressed in Escherichia coli and purified for biochemical characterization. The enzyme was shown to act as an endo-1,4-β-xylanase, but minor activities
against lichenan, barley glucan, methylumbelliferyl cellobioside and p-nitrophenyl xyloside were also detected. The specific activity and pH and temperature optima for hydrolysis of oat xylan
were 111.3 U⋅mg-1, 5.5 and 102°C, respectively. The endoxylanase was stable at 90°C and retained 50% activity when incubated for 2 h at 100°C.
Received: 19 May 1995/Received revision: 31 July 1995/Accepted: 7 September 1995 相似文献
12.
A method for reducing RNA and DNA in the bacterium Methylococcus capsulatus (Bath) has been developed. Endogenous RNase and DNase were activated by a 10 s heat shock at 90°C. Cells were then incubated
at 60°C for 20 min to allow degradation of the nucleic acids. The optimum pH for the process was 7.0. The protein loss was
less than 10% and occurred during the initial heat shock. No protein loss was found during incubation. The total dry-weight
loss in connection with an 80% reduction of the nucleic acid content was 20%–25%, giving a final product with a raw protein
content of approximately 75%. Reduction of both RNA and DNA was inhibited by CuSO4 and ZnSO4. DNA reduction was stimulated by other minerals. Optimal stimulation was found at 1 mM FeSO4. Reduction of RNA was not increased by any of the minerals tested.
Received: 29 June 1995/Received last revision: 2 October 1995/Accepted: 16 October 1995 相似文献
13.
G. R. V. Babu O. K. Vijaya V. L. Ross J. H. Wolfram K. D. Chapatwala 《Applied microbiology and biotechnology》1996,45(1-2):273-277
Our isolate, Pseudomonas putida, is known to be capable of utilizing cyanides as the sole source of carbon (C) and nitrogen (N) both in the form of free cells
and cells immobilized in calcium alginate. In the present study, the cell-free extract(s) were prepared from the cells of
P. putida grown in the presence of sodium cyanide. The ability of enzyme(s) to convert cyanides, cyanates, thiocyanates, formamide and
cyanide-containing mine waters into ammonia (NH3) was studied at pH 7.5 and pH 9.5. The kinetic analysis of cyanide and formamide conversion into NH3 at pH 7.5 and pH 9.5 by the cell-free extract(s) of P. putida was also studied. The K
m and V
max values for cyanide/formamide were found to be 4.3/8 mM and 142/227 μmol NH3 released mg protein-1 min-1 respectively at pH 7.5 and 5/16.67 mM and 181/434 μmol NH3 released mg protein-1 h-1 respectively at pH 9.5. The study thus concludes that the cell-free extract(s) of P. putida is able to metabolize not only cyanides, cyanates, thiocyanates, and formamide but also cyanide-containing mine waters to
NH3.
Received: 10 April 1995/Received revision: 24 July 1995/Accepted: 22 August 1995 相似文献
14.
Keratinase of Doratomyces microsporus 总被引:10,自引:0,他引:10
The fungus Doratomyces microsporus produced an extracellular keratinase during submerged aerobic cultivation in a medium containing a protein inducer for enzyme
synthesis. The keratinase was purified to homogeneity using hydrophobic interaction chromatography followed by gel chromatography.
The molecular weight was estimated to be 33 kDa (from SDS-PAGE analysis) or 30 kDa (by gel chromatography), suggesting a monomeric
structure. The isoelectric point of the enzyme was determined to be around 9. The optimal pH and temperature for keratinolytic
activity were pH 8–9 and 50 °C, respectively. The serine protease inhibitor PMSF totally inhibited the keratinase. The enzyme
was not glycosylated. It was capable of hydrolysing different keratinous materials as well as some non-keratinous proteins.
Hydrolysis of some synthetic substrates, specific for known proteinases, suggested that the keratinase of D. microsporus is close to proteinase K.
Received: 9 July 1999 / Received revision: 13 September 1999 / Accepted: 17 September 1999 相似文献
15.
J. Sánchez-Torres P. Pérez R. I. Santamaría 《Applied microbiology and biotechnology》1996,46(2):149-155
Several alkalophilic Bacillus spp. strains were selected for their capacity to produce alkaline cellulases. Culture supernatants of these strains showed
optimal cellulase activities between pH 8 and 9 and they were stable from pH 6 to pH 12. A cellulase gene (celB1) from the alkalophilic Bacillus sp. strain N186-1 was cloned in Escherichia coli using polymerase chain reaction techniques. The cloned gene was present in a 2.539-bp HindIII fragment and its nucleotide sequence was determined. The coding sequence showed an open-reading frame encoding 389 amino
acids. The amino acid sequence, deduced from the nucleotide sequence, permitted us to include it in family 5 (or A) of the
glycosyl hydrolases. The complete open-reading frame of celB1 was cloned in the plasmid pET-11d and expressed in E. coli BL21 (DE3), in which a protein of 39 kDa was obtained in the cytoplasm; however, no endoglucanase activity was detected.
A second construction in pET-12a allowed the production of a 39-kDa protein located in the periplasmic space of E. coli that had endoglucanase activity. The protein produced has optimal activity at pH 7 and 50°C and it retains more than 70%
of its activity after incubation for 1 h at pH 12.
Received: 27 December 1995/Received revision: 14 March 1996/Accepted: 25 March 1996 相似文献
16.
The effects of fermentation of cassava by Aspergillus niger B-1 β-glucosidase on its cyanide and protein content, and the optimal conditions for this enzyme’s activity, were examined.
This fermentation process reduced the cyanide content of cassava by 95% to 2 mg/kg, and increased its total protein content
by 50%, thereby improving its nutritional value. A significant decrease in cyanogenic glycosides was detected after 3 days
of fermentation. The optimal pH for A. nigerβ-glucosidase activity on the cyanogenic glycoside linamarin was determined to be 3, the optimal temperature 55 °C, and its
K
m 0.3 mM. The findings presented here will facilitate the development of an improved method for detoxification of cassava and
for enhancement of its nutritional value.
Received: 17 August 1995/Received revision: 27 October 1995/Accepted: 30 October 1995 相似文献
17.
A Bayoudh N Gharsallah M Chamkha A Dhouib S Ammar M Nasri 《Journal of industrial microbiology & biotechnology》2000,24(4):291-295
An alkaline protease produced by Pseudomonas aeruginosa MN1, isolated from an alkaline tannery waste water, was purified and characterized. The enzyme was purified 25-fold by gel
filtration and ion exchange chromatography to a specific activity of 82350 U mg−1. The molecular weight of the enzyme was estimated to be 32000 daltons. The optimum pH and temperature for the proteolytic
activity were pH 8.00 and 60°C, respectively. Enzyme activity was inhibited by EDTA suggesting that the preparation contains
a metalloprotease. Enzyme activity was strongly inhibited by Zn2+, Cu2+ and Hg2+(5 mM), while Ca2+ and Mn2+ resulted in partial inhibition. The enzyme is different from other Pseudomonas aeruginosa alkaline proteases in its stability at high temperature; it retained more than 90% and 66% of the initial activity after
15 and 120 min incubation at 60°C. Journal of Industrial Microbiology & Biotechnology (2000) 24, 291–295.
Received 09 June 1999/ Accepted in revised form 24 January 2000 相似文献
18.
A. Hartke A. Benachour P. Boutibonnes Y. Auffray 《Applied microbiology and biotechnology》1996,45(1-2):132-136
Two type-II restriction endonucleases, BloI and BloII, have been detected in a Bifidobacterium longum strain. BloI is influenced by dam methylation: it cleaves dam
- but not dam
+ DNA. It shows a temperature and pH optimum of 45°C and pH 7.5. Restriction analysis and cloning experiments showed that the
recognition sequence is RGATCY and that the enzyme cuts 5′ to the guanine residue. It is an isoschizomer of commercial enzymes,
BstYI and XhoII. The second activity is not inhibited by dam methylation. It has a temperature optimum between 25°C and 30°C and shows a broad pH optimum between 4.5 and 7.0. The activity
is thermolabile and can be heat-killed by a 5 min incubation at 60°C. Cloning and sequencing experiments revealed that its
recognition sequence is CTGCAG and that it cuts 5′ to the second guanine residue in the sequence. This enzyme is the first
described isoschizomer of PstI.
Received: 22 May 1995/Accepted: 26 July 1995 相似文献
19.
A. De Smul J. Dries L. Goethals H. Grootaerd W. Verstraete 《Applied microbiology and biotechnology》1997,48(3):297-303
In a mesophilic (30–35 °C), sulphidogenic, ethanol-fed expanded-granular-sludge-blanket reactor, sulphate, at loading rates
of up to 10.0–12.0 g Sl−1␣day−1, was removed with an average efficiency of more than 80%. The pH was between 7.7 and 8.3 and the maximal total dissolved
sulphide concentration was up to 20 mM S (650 mg S/l). The alkaline pH was maintained by either a pH-control unit with sodium
hydroxide or by stripping part of the sulphide and CO2 from the recycle with nitrogen gas. The superficial upstream liquid velocity (v
up) was 3.0–4.5 m/h. The ratio of ethanol to sulphur was near stoichiometry. At alkaline pH, the activity of the acetotrophic
sulphate-reducing bacteria, growing on acetate, was strongly enhanced, whereas at pH below 7.7 the acetotrophic sulphate-reducing
bacteria were inhibited by aqueous H2S. With regard to the removal efficiency and operational stability, external stripping with N2 and pH control were equally successful.
Received: 2 December 1996 / Received revision: 13 March 1997 / Accepted: 15 March 1997 相似文献
20.
Studies were carried out to utilize in situ proteases of shrimp heads to recover carotenoproteins possessing antioxidant activity.
Highest protease activity of the buffer extract was found at pH 8.0 (9.85 ± 0.61 units). The protease activity increased with
temperature up to 50°C and reduced thereafter with highest activity being 19.32 ± 2.0 units. Thus, the autolysis of shrimp
heads for recovery of carotenoprotein was carried out at pH 8.0 and at 50°C. Waste to buffer ratio had a significant (p < 0.05)
effect on recovery of carotenoids in carotenoprotein filtrate with a maximum of 58.5 ± 6.4% recovery with a waste to buffer
ratio of 1:2.5 (w:v). The carotenoid recovery increased significantly to 63.4% ± 3.6% at the end of a 4-h autolysis. The studies on combined
effect of waste to buffer ratio and autolysis time indicated increase in protein recovery with increase in waste to buffer
ratio but not with autolysis time. DPPH scavenging activity of the carotenoprotein isolate increased with autolysis time up
to 100 min, and thereafter, reduced above 160 min of autolysis time. With increase in waste to buffer ratio, the scavenging
activity increased, reaching more than 12.5 mg TBHQ equivalent/mg protein at waste to buffer ratio of 1:5. The optimum autolysis
condition for obtaining antioxidant activity rich carotenoprotein from shrimp heads was found to be waste to buffer (pH 8.0)
ratio of 1:5 and an autolysis time of 2 h at 50°C. The isolated carotenoprotein was found to have antioxidant activity with
respect to singlet oxygen quenching, reducing power and metal chelating activity. 相似文献