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1.
Determination of the apparent pK
a's of purified carboxymethylcellulases fromAspergillus niger andCellulomonas biazotea at different temperatures and in the presence of dioxane indicated two side chain carboxyl groups which controlled the limiting
rate in both organisms. The thermostability of both enzymes slightly decreased with increasing pH from 5 to 7.5 but was unaffected
in the presence of 0.5 mmol/L Mn2+. The CMCase fromC. biazotea had an activation energy of 35 kJ/mol and a half-life of 89 min in the presence of 8 mol/L urea at 40°C. The half-life of
CMCase fromA. niger in 8 mol/L urea and at 37°C was 125 min as determined by a 0–9 mol/L transverse urea gradient PAGE. The CMCases fromA. niger andC. biazotea had the same thermostabilities in the absence of CMC although the enzyme from the former was more thermostable in the presence
of the substrate. The CMCase fromA. niger was also more efficient in hydrolyzing CMC than the enzyme fromC. biazotea. 相似文献
2.
The activity of glutamine synthetase fromAspergillus niger was significantly lowered under conditions of citric acid fermentation. The intracellular pH of the organism as determined
by bromophenol blue dye distribution and fluorescein diacetate uptake methods was relatively constant between 6·0–6·5, when
the pH of the external medium was varied between 2·3–7·0.Aspergillus niger glutamine synthetase was rapidly inactivated under acidic pH conditions and Mn2+ ions partially protected the enzyme against this inactivation. Mn2+-dependent glutamine synthetase activity was higher at acidic pH (6·0) compared to Mg2+-supported activity. While the concentration of Mg2+ required to optimally activate glutamine synthetase at pH 6·0 was very high (≥ 50 mM), Mn2+ was effective at 4 mM. Higher concentrations of Mn2+ were inhibitory. The inhibition of both Mn2+ and Mg2+-dependent reactions by citrate, 2-oxoglutarate and ATP were probably due to their ability to chelate divalent ions rather
than as regulatory molecules. This suggestion was supported by the observation that a metal ion chelator, EDTA also produced
similar effects. Of the end-products of the pathway, only histidine, carbamyl phosphate, AMP and ADP inhibitedAspergillus niger glutamine synthetase. The inhibitions were more pronounced when Mn2+ was the metal ion activator and greater inhibition was observed at lower pH values. These results permit us to postulate
that glutamine synthesis may be markedly inhibited when the fungus is grown under conditions suitable for citric acid production
and this block may result in delinking carbon and nitrogen metabolism leading to acidogenesis 相似文献
3.
The biochemical rationale for the inhibition of citric acid fermentation by Aspergillus niger in the presence of Mn2+ ions has been investigated using high citric acid-yielding, Mn2+ ion-sensitive as well as Mn2+ ion-tolerant mutant strains of A. niger. In the presence of Mn2+ (1.5 mg/l), citric acid production by the Mn2+ ion-sensitive strain (KCU 520) was reduced by about 75% with no apparent effect on citric acid yield by the Mn2+ ion-tolerant mutant strain (GS-III) of A. niger. The significantly increased level of the Mn2+ ion-requiring NADP+-isocitrate dehydrogenase activity in KCU 520 cells and the lack of effect on the activity level of the enzyme in GS-III mutant cells by Mn2+ ions during fermentation seem to be responsible for the Mn2+ ion inhibition of citric acid production by the KCU 520 strain and the high citric acid yield by the mutant strain GS-III of A. niger even in the presence of Mn2+. 相似文献
4.
The specific activity of glutamine synthetase (L-glutamate: ammonia ligase, EC 6.3.1.2) in surface grownAspergillus niger was increased 3–5 fold when grown on L-glutamate or potassium nitrate, compared to the activity obtained on ammonium chloride.
The levels of glutamine synthetase was regulated by the availability of nitrogen source like NH
4
+
, and further, the enzyme is repressed by increasing concentrations of NH
4
+
. In contrast to other micro-organisms, theAspergillus niger enzyme was neither specifically inactivated by NH
4
+
or
L-glutamine nor regulated by covalent modification. Glutamine synthetase fromAspergillus niger was purified to homogenity. The native enzyme is octameric with a molecular weight of 385,000±25,000. The enzyme also catalyses
Mn2+ or Mg2+-dependent synthetase and Mn2+-dependent transferase activity.
Aspergillusniger glutamine synthetase was completely inactivated by two mol of phenyl-glyoxal and one mol of N-ethylmaleimide with second
order rate constants of 3.8 M-1 min-1 and 760 M-1 min-1 respectively. Ligands like Mg. ATP, Mg. ADP, Mg. AMP, L-glutamate NH
4
+
, Mn2+ protected the enzyme against inactivation. The pattern of inactivation and protection afforded by different ligands against
N-ethylamaleimide and phenylglyoxal was remarkably similar. These results suggest that metal ATP complex acts as a substrate
and interacts with an arginine ressidue at the active site. Further, the metal ion and the free nucleotide probably interact
at other sites on the enzyme affecting the catalytic activity. 相似文献
5.
The present work reports for the first time the purification and characterisation of two extremely halotolerant endo-xylanases from a novel halophilic bacterium, strain CL8. Purification of the two xylanases, Xyl 1 and 2, was achieved by anion exchange and hydrophobic interaction chromatography. The enzymes had relative molecular masses of 43 kDa and 62 kDa and pI of 5.0 and 3.4 respectively. Stimulation of activity by Ca2+, Mn2+, Mg2+, Ba2+, Li2+, NaN3 and isopropanol was observed. The Km and Vmax values determined for Xyl 1 with 4-O-methyl-d-glucuronoxylan are 5 mg/ml and 125,000 nkat/mg respectively. The corresponding values for Xyl 2 were 1 mg/ml and 143,000 nkat/mg protein. Xylobiose and xylotriose were the major end products for both endoxylanases. The xylanases were stable at pH 4–11 showing pH optima around pH 6. Xyl 1 shows maximal activity at 60°C, Xyl 2 at 65°C (at 4 M NaCl). The xylanases showed high temperature stability with half-lives at 60°C of 97 min and 192 min respectively. Both xylanases showed optimal activity at 1 M NaCl, but substantial activity remained for both enzymes at 5 M NaCl.Communicated by W.D. Grant 相似文献
6.
A. BarbosaJr L. H. S. GuimarÃes H. F. Terenzi J. A. Jorge F. A. Leone M. L. T. M. Polizeli 《Folia microbiologica》2008,53(6):509-516
The biochemical properties of the alkaline phosphatases (AlPs) produced by Rhizopus microsporus are described. High enzymic levels were produced within 1–2 d in agitated cultures with 1 % wheat bran. Intra- and extracellular
AlPs were purified 5.0 and 9.3×, respectively, by DEAE-cellulose and ConA-sepharose chromatography. Molar mass of 118 and
120 kDa was estimated by gel filtration for both forms of phosphatases. SDS-PAGE indicated dimeric structures of 57 kDa for
both forms. Mn2+, Na+ and Mg2+ stimulated the activity, while Al3+ and Zn2+ activated only the extracellular form. Optimum temperature and pH for both phosphatases were 65 °C and pH 8.0, respectively.
The enzymes were stable at 50 °C for at least 15 min. Hydrolysis of 4-nitrophenyl phosphate exhibited a K
m 0.28 and 0.22 mmol/L, with υ
lim 5.89 and 4.84 U/mg, for intra- and extracellular phosphatases, respectively. The properties of the reported AlPs may be suitable
for biotechnological application. 相似文献
7.
Purification and properties of an intracellular leucine aminopeptidase from the fungus,Penicillium citrinum strain IFO 6352 总被引:1,自引:0,他引:1
An intracellular leucine aminopeptidase (LAP) fromPenicillium citrinum (IFO 6352) was purified to homogeneity using three successive purification steps. The enzyme has a native molecular mass of 63 kDa using HPLC gel filtration analysis and a molecular mass of 65 kDa when using SDS-polyacrylamide gel electrophoresis. This monomeric aminopeptidase showed maximum enzyme activity at pH 8.5. An optimum temperature was 45–50°C whenl-Leu-p-nitroanilide (pNA) was the substrate, and enzyme activity drastically decreased above 60°C. The Michaelis-Menten constants forl-Leu-pNA andl-Met-pNA were 2.7 mM and 1.8 mM, respectively. When the enzyme reacted with biosynthetic methionyl human growth hormone, it showed high specificity for N-terminal methionine residue and recognized a stop sequence (Xaa-Pro). The aminopeptidase was inactivated by EDTA or 1,10-phenanthroline, indicating that it is a metallo-exoprotease. Enzyme activity was restored to 90% of maximal activity by addition of Co2+ ions. The activity of EDTA-treated enzyme was restored by addition of Zn2+, but reconstitution with Ca2+, Mg2+ or Mn2+ restored some enzyme activity. It is likely that Co2+ ions play an important role in the catalysis or stability of thePenicillium citrinum aminopeptidase, as zinc plays a similar function in other leucine aminopeptidases. 相似文献
8.
The extracellular activity ofAspergillus niger phytase at the end of the growth phase was 132 nkat/mL in a laboratory bioreactor. The purified enzyme has molar mass approximately
100 kDa, pH optimum at 5.0, temperature optimum at 55°C and high pH and temperature stability. TheK
m for dodecasodium phytate, calcium phytate and 4-nitrophenyl phosphate are 0.44, 0.45 and 1.38 mmol/L, respectively. The enzyme
is noncompetively inhibited by inorganic monophosphate (K
i=2.85 mmol/L) and by Cu2+, Zn2+, Hg2+, Sn2+, Cd2+ ions and strongly by F− ones; it is activated by Ca2+, Mg2+ and Mn2+ ions. The substrate specificity of phytase is broad with the highest affinity to calcium phytate. 相似文献
9.
Streptomycetes have two distinct glutamine synthetases (GS): a heat-stable dodecameric GSI and a heat-labile octameric GSII.
A heat-inactivated GS activity was detected in crude extracts ofStreptomyces cinnamonensis cells grown with nitrate or glutamate as the nitrogen source. The purified enzyme obtained from crude extracts of the nitrate-grown
cells after affinity and anion-exchange chromatography was also heat-labile; it was inactivated by 80 % when incubated at
50 °C for 1 h. However, the enzyme has properties typical of GSI and similar with those of the heat-stable GSI purified fromS. aureofaciens: It is composed of twelve subunits, each ofM 55 kDa, and has a native molar mass of 625 kDa and an isoelectric point at pH 4.2. In addition, its activity is regulated
by reversible adenylylation. Mg2+ and NaCl but not Mn2+ protected the purified enzyme from thermal inactivation, and both NaCl and Mn2+ or Mg2+ stabilized its activity at 4–8 °C. As compared with GSI fromS. aureofaciens, theS. cinnamonensis enzyme was cleaved more extensively during SDS-PAGE, was less sensitive to feedback inhibitors, and similarly affected by
divalent cations. TheK
m values were 12.5 mmol/L forl-glutamate, 0.1 for NH
4
+
, 1.25 for ATP, 18.5 forl-glutamine, 3.3 for hydroxylamine and 0.087 for ADP. To our best knowledge, this is the first report of a heatlabile GSI from
any source. 相似文献
10.
Sarvesh Kumar Soni Jayant Malhar Khire 《World journal of microbiology & biotechnology》2007,23(11):1585-1593
Novel extracellular phytase was produced by Aspergillus niger NCIM 563 under submerged fermentation conditions at 30 °C in medium containing dextrin and glucose as carbon sources along
with sodium nitrate as nitrogen source. Maximum phytase activity (41.47 IU/mL at pH 2.5 and 10.71 IU/mL at pH 4.0) was obtained
when dextrin was used as carbon source along with glucose and sodium nitrate as nitrogen source. Nearly 13 times increase
in phytase activity was observed when phosphate in the form of KH2PO4 (0.004 g/100 mL) was added in the fermentation medium. Physic-chemical properties of partially purified enzyme indicate the
possibility of two distinct forms of phytases, Phy I and Phy II. Optimum pH and temperature for Phy I was 2.5 and 60 °C while
Phy II was 4.0 and 60 °C, respectively. Phy I was stable in the pH range 1.5–3.5 while Phy II was stable in the wider pH range,
2.0–7.0. Molecular weight of Phy I and Phy II on Sephacryl S-200 was approximately 304 kDa and 183 kDa, respectively. Phy
I activity was moderately stimulated in the presence of 1 mM Mg2+, Mn2+, Ca2+ and Fe3+ ions and inhibited by Zn2+ and Cd2+ ions while Phy II activity was moderately stimulated by Fe3+ ions and was inhibited by Hg2+, Mn2+ and Zn2+ ions at 1 mM concentration in reaction mixture. The Km for Phy I and II was 3.18 and 0.514 mM while Vmax was 331.16 and 59.47 μmols/min/mg protein, respectively. 相似文献
11.
The high-molar mass from of β-glucosidase fromAspergillus niger strain NIAB280 was purified to homogeneity with a 46-fold increase in purification by a combination of ammonium sulfate precipitation,
hydrophobic interaction, ion-exchange and gel-filtration chromatography. The native and subunit molar mass was 330 and 110
kDa, respectively. The pH and temperature optima were 4.6–5.3 and 70°C, respectively. TheK
m andk
cat for 4-nitrophenyl β-d-glucopyranoside at 40°C and pH 5 were 1.11 mmol/L and 4000/min, respectively. The enzyme was activated by low and inhibited
by high concentrations of NaCl. Ammonium sulfate inhibited the enzyme. Thermolysin periodically inhibited and activated the
enzyme during the course of reaction and after 150 min of proteinase treatment only 10% activity was lost with concomitant
degradation of the enzyme into ten low-molar-mass active bands. When subjected to 0–9 mol/L transverse urea-gradient-PAGE
for 105 min at 12°C, the nonpurified β-glucosidase showed two major bands which denatured at 4 and 8 mol/L urea, respectively,
with half-lives of 73 min. 相似文献
12.
Wang SL Lin YT Liang TW Chio SH Ming LJ Wu PC 《Journal of industrial microbiology & biotechnology》2009,36(1):65-73
A lipase-producing bacterium was isolated and identified as Pseudomonas monteilii TKU009. A lipase (F2) and lipase-like materials (F1) were purified from the culture supernatant of P. monteilii TKU009 with soybean powder as the sole carbon/nitrogen source. The molecular mass of F1 and F2 was estimated to be 44 kDa
by SDS-PAGE and gel filtration. The optimum pH, optimum temperature, and pH and thermal stabilities of F2 were 7, 40°C, 8–11,
and 50°C; and of F1 were 6, 40°C, 6–7, and 50°C, respectively. F2 was completely inhibited by EDTA and slightly by Mg2+, Fe2+, Mn2+, and SDS. F1 was completely inhibited by EDTA and Fe2+ and strongly by Zn2+, Mn2+, Ca2+, Mg2+, and SDS. The activities of both the enzymes were enhanced by the addition of non-ionic surfactants Triton X–100 and Tween
40, especially for F1. F2 preferably acted on substrates with a long chain (C10–C18) of fatty acids, while F1 showed a broad
spectrum on those with chain length of C4–C18. The marked activity of F2 in organic solvents makes it an ideal choice for
application in a water-restricted medium including organic synthesis.
Li-June Ming is a visiting Professor at the National Cheng Kung University. 相似文献
13.
Natural and recombinant fungal laccases for paper pulp bleaching 总被引:10,自引:0,他引:10
Sigoillot C Record E Belle V Robert JL Levasseur A Punt PJ van den Hondel CA Fournel A Sigoillot JC Asther M 《Applied microbiology and biotechnology》2004,64(3):346-352
Three laccases, a natural form and two recombinant forms obtained from two different expression hosts, were characterized and compared for paper pulp bleaching. Laccase from Pycnoporus cinnabarinus, a well known lignolytic fungus, was selected as a reference for this study. The corresponding recombinant laccases were produced in Aspergillus oryzae and A. niger hosts using the lacI gene from P. cinnabarinus to develop a production process without using the expensive laccase inducers required by the native source. In flasks, production of recombinant enzymes by Aspergilli strains gave yields close to 80 mg l–1. Each protein was purified to homogeneity and characterized, demonstrating that the three hosts produced proteins with similar physico-chemical properties, including electron paramagnetic resonance spectra and N-terminal sequences. However, the recombinant laccases have higher Michaelian (K
m) constants, suggesting a decrease in substrate/enzyme affinity in comparison with the natural enzyme. Moreover, the natural laccase exhibited a higher redox potential (around 810 mV), compared with A. niger (760 mV) and A. oryzae (735 mV). Treatment of wheat straw Kraft pulp using laccases expressed in P. cinnabarinus or A. niger with 1-hydroxybenzotriazole as redox mediator achieved a delignification close to 75%, whereas the recombinant laccase from A. oryzae was not able to delignify pulp. These results were confirmed by thioacidolysis. Kinetic and redox potential data and pulp bleaching results were consistent, suggesting that the three enzymes are different and each fungal strain introduces differences during protein processing (folding and/or glycosylation). 相似文献
14.
Five commercial preparations of glucoamylases (three fromAspergillus niger, one each fromAspergillus foetidus andAspergillus candidus) were purified by ultrafiltration, Sepharose-gel filtration and DEAE-sephadex chromatography. Two forms of the enzyme, namely
glucoamylase I and glucoamylase II were obtained from the fungi except from one strain ofA. Niger. All the enzymes appeared homogeneous by electrophoresis and ultracentrifugation. The specific activities varied between 85
and 142 units. The pH and temperature optima were between 4 and 5, and 60‡C respectively. The molecular weight as determined
by the sodium dodecyl sulphate-polyacrylamide gel electrophoresis ranged from 75,000 to 79,000 for glucoamylase I and 60,000
to 72,000 for glucoamylase II. OnlyA. niger glucoamylases contained phenylalanine at the N-terminal end. The amino acid composition of the enzymes was generally similar.
However,A. niger andA. foetidus glucoamylases, in contrast toA. candidus enzymes, contained greater percentage of acidic than of basic amino acids. The enzymes contained 15 to 30% carbohydrate and
49 to 57 residues of monosaccharides per mol.A. niger enzymes contained mannose, glucose, galactose, xylose and glucosamine but theA. candidus enzyme lacked xylose and glucose and only xylose was absent inA, foetidus enzymes. Majority of the carbohydrate moieties were O-glycosidically linked through mannose to the hydroxyl groups of seline
and threonine of the polypeptide chain. 相似文献
15.
L B Crotti H F Terenzi J A Jorge M L T M Polizeli 《Journal of industrial microbiology & biotechnology》1998,20(3-4):238-243
Pectolytic enzymes from the hyperproducer exo-1 mutant of Neurospora crassa are induced either by pectin or galactose. Galactose-induced pectinases, in contrast with pectin-induced enzymes, are not
affected by glucose repression. Here, the pectolytic enzymes induced by galactose were purified and characterized. Extracellular
pectolytic activities were separated into two main fractions. Pool I contained lyases, and a polygalacturonase (PG) copurifying
as a complex of about 80 kDa (gel filtration). Pool II contained PG only. Under urea-SDS-PAGE the lyases and polygalacturonase
from pool I migrated with an apparent MW of 56.2 kDa, and 34.3 kDa, respectively. PG from pool II exhibited an apparent MW
of 44.7 kDa. Cell extracts contained PG free of lyase activities. Purified intracellular PG migrated (SDS-PAGE) as a single
band of apparent MW of 31.5 kDa. All pectinases were glycoproteins (18.5–39% carbohydrate), with stability and optimum pH
at 5–6 and 9–10 for PG and lyases, respectively. Temperature optima were 40–50°C, respectively. All enzymes were inactivated
at 60°C, with a half-life from 1.5 to 5 min. Activation energy (Ea) values for extracellular and intracellular PG varied between
0.45 and 2.0 Kcal mol−1. Pool II and intracellular PG and lyases, exhibited a random mechanism of hydrolysis. Pool I PG exhibited an exo character.
Received 20 October 1997/ Accepted in revised form 28 February 1998 相似文献
16.
A study was carried out on the application of supercritical fluid to the hydrolysis of boll fibers of cotton (cultivar Tashkent-6
ofGossypium hirsutum L.) by cellulase enzymes fromTrichoderma viride, Trichoderma reesei andAspergillus niger. Conditions of the enzymatic process were optimized. The stabilities of cellulase enzymes were sustained, at the pressure
of up to 160 atm for 48 hours at 50°C in supercritical carbon dioxide. 相似文献
17.
Barnali Ray Basu Ajit K. Banik Manas Das 《World journal of microbiology & biotechnology》2008,24(4):449-455
Fish scale, the chief waste material of fish processing industries was processed and tested for production of extracellular
protease by mutant Aspergillus niger AB100. Protease production by A. niger AB100 was greatly enhanced in presence of processed fish scale powder. Where as among the three complex nutrients tested, soya
bean meal shows maximum stimulatory effect over protease production (2,776 μmol/ml/min) when used in combination with glucose
(5% w/v) and urea (2.5% w/v). The protease was optimally active at pH 7.0, retaining more than 60% of its activity in the
pH range of 5–9. The enzyme was found to be most active at 50°C and stable at 30°C for 1 h. Purification of enzyme by CM-Cellulose
and SDS-PAGE resulted in about 26-fold increase in the specific activity of the enzyme with a molecular weight of 30.9 kDa.
HPLC study shows the purity of the enzyme as 75.92%. By the activating effect of divalent cations (Fe2+, Zn2+, Mn2+, Ca2+and Mg2+) and inhibiting effect of chelating agent (EDTA) and Hg2+, the enzyme was found to be a metalloprotease. 相似文献
18.
The class II fructose-1,6-bisphosphatase gene of Corynebacterium glutamicum, fbp, was cloned and expressed with a N-terminal His-tag in Escherichia coli. Purified, His-tagged fructose-1,6-bisphosphatase from C. glutamicum was shown to be tetrameric, with a molecular mass of about 140 kDa for the homotetramer. The enzyme displayed Michaelis-Menten kinetics for the substrate fructose 1,6-bisphosphate with a Km value of about 14 µM and a Vmax of about 5.4 µmol min–1 mg–1 and kcat of about 3.2 s–1. Fructose-1,6-bisphosphatase activity was dependent on the divalent cations Mg2+ or Mn2+ and was inhibited by the monovalent cation Li+ with an inhibition constant of 140 µM. Fructose 6-phosphate, glycerol 3-phosphate, ribulose 1,5-bisphosphate and myo-inositol-monophosphate were not significant substrates of fructose-1,6-bisphosphatase from C. glutamicum. The enzymatic activity was inhibited by AMP and phosphoenolpyruvate and to a lesser extent by phosphate, fructose 6-phosphate, fructose 2,6-bisphosphate, and UDP. Fructose-1,6-bisphosphatase activities and protein levels varied little with respect to the carbon source. Deletion of the chromosomal fbp gene led to the absence of any detectable fructose-1,6-bisphosphatase activity in crude extracts of C. glutamicum WTfbp and to an inability of this strain to grow on the carbon sources acetate, citrate, glutamate, and lactate. Thus, fbp is essential for growth on gluconeogenic carbon sources and likely codes for the only fructose-1,6-bisphosphatase in C. glutamicum. 相似文献
19.
Agathe Garnier Ali Berredjem Bernard Botton 《Fungal genetics and biology : FG & B》1997,22(3):168-176
The NAD-dependent glutamate dehydrogenase (GDH) (EC 1.4.1.2) fromLaccaria bicolorwas purified 410-fold to apparent electrophoretic homogeneity with a 40% recovery through a three-step procedure involving ammonium sulfate precipitation, anion-exchange chromatography on DEAE–Trisacryl, and gel filtration. The molecular weight of the native enzyme determined by gel filtration was 470 kDa, whereas sodium dodecyl sulfate–polyacrylamide gel electrophoresis gave rise to a single band of 116 kDa, suggesting that the enzyme is composed of four identical subunits. The enzyme was specific for NAD(H). The pH optima were 7.4 and 8.8 for the amination and deamination reactions, respectively. The enzyme was found to be highly unstable, with virtually no activity after 20 days at −75°C, 4 days at 4°C, and 1 h at 50°C. The addition of ammonium sulfate improved greatly the stability of the enzyme and full activity was still observed after several months at −75°C. NAD-GDH activity was stimulated by Ca2+and Mg2+but strongly inhibited by Cu2+and slightly by the nucleotides AMP, ADP, and ATP. The Michaelis constants for NAD, NADH, 2-oxoglutarate, and ammonium were 282 μM, 89 μM, 1.35 mM, and 37 mM, respectively. The enzyme had a negative cooperativity for glutamate (Hill number of 0.3), and itsKmvalue increased from 0.24 to 3.6 mM when the glutamate concentration exceeded 1 mM. These affinity constants of the substrates, compared with those of the NADP-GDH of the fungus, suggest that the NAD-GDH is mainly involved in the catabolism of glutamate, while the NADP-GDH is involved in the catalysis of this amino acid. 相似文献
20.
A Novel Ferric Reductase Purified from <Emphasis Type="Italic">Magnetospirillum gryphiswaldense</Emphasis> MSR-1 总被引:1,自引:0,他引:1
A ferric reductase was purified into an electrophoretically homologous state from Magnetospirillum gryphiswaldense MSR-1 strain. The enzyme was found within the cytoplasm and associated with the cytoplasmic membrane. The molecular weight
of the purified enzyme was calculated as 16.1 kDa using sodium dodecyl sulfate–polyacrylamide gel electrophoresis, and was
almost identical to that calibrated using agarose gel filtration chromatography. It was NADH-dependent and required flavin
mononucleotide as a cofactor. The optimal reaction temperature and pH values were 30°C and 6.5, respectively. The K
m and Vmax values for ferric citrate were 45.1 μM and 1.216 μM min−1, respectively. Though ferric reductase activity could be inhibited by Co2+, Cu2+, Mn2+, and Zn2+, even high concentrations of Mg2+ ions have failed to accomplish such enzyme inhibition. Furthermore, the molecular weight, the N-terminal sequence, and the
activity of ferric reductase from MSR-1 are not matching with the enzyme preparation obtained from an analogous strain M. magnetotacticum (MS-1). Therefore, it is concluded that the ferric reductase of M. grysphiwaldense and M. magnetotacticum strains are two different enzymes. 相似文献