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1.
Jeyaraman Vikramathithan Gali Nirmal kumar Pandurangan Muthuraman Kotteazeth Srikumar 《The protein journal》2010,29(7):481-486
A thermo stable xylanase was purified and characterized from the cladodes of Cereus pterogonus plant species. The enzyme was purified to homogeneity by ammonium sulfate (80%) fractionation, ion exchange and size exclusion
chromatography. The enzyme showed a final specific activity of 216.2 U/mg and the molecular mass of the protein was 80 KDa.
The optimum pH and temperature for xylanase activity were 5.0 and 80 °C, respectively,. With oat spelt xylan as a substrate
the enzyme yielded a Km value of 2.24 mg/mL and a Vmax of 5.8 μmol min−1 mg−1. In the presence of metal ions (1 mM) such as Co2+,Mn2+, Ni2+, Ca2+ and Fe3+ the activity of the enzyme increased, where as strong inhibition of the enzyme activity was observed with the use of Hg2+, Cd2+, Cu2+, while partial inhibition was noted with Zn2+ and Mg2+. The substrate specificity of the xylanase yielded maximum activity with oat spelt xylan. 相似文献
2.
Mohsen Mohamed Selim Asker Mohamed Fawzy Ramadan Samir Khalf Abd El-Aal Ebtsam Mokhtar Mohamed El-Kady 《World journal of microbiology & biotechnology》2009,25(5):789-794
Trehalose synthase (TSII) from Corynebacterium nitrilophilus NRC was successively purified by ammonium sulphate precipitation, ion exchange chromatography on DEAE-cellulose and gel filtration
chromatography on Sephadex G-100 columns. The specific activity of the trehalose synthase was increased ~200-fold, from 0.14
U mg−1 protein to 28.3 U mg−1 protein. TSII was found to be a monomeric protein with a molecular weight of 67–69 kDa. Characterization of the enzyme exhibited
optimum pH and temperature were 7.5 and 35°C, respectively. The purified enzyme was stable from pH 6.6 to 7.8 and able to
prolong its thermal stability up to 35°C. The enzyme activity was inhibited strongly by Zn2+, Hg2+ and Cu2+ and moderately by Ba2+, Fe2+, Pb2+ and Ni2+. Other metal ions Ca2+, Mg2+, Co2+, Mn2+ and EDTA had almost no effect. 相似文献
3.
Complexes formed by reduced glutathione (GSH) with metal cations (Cr2+, Mn2+,Fe2+,Co2+,Ni2+,Cu2+,Zn2+,Cd2+,Hg2+) were systematically investigated by the density functional theory (DFT). The results showed that the interactions of the metal cations with GSH resulted in nine different stable complexes and many factors had an effect on the binding energy. Generally, for the same period of metal ions, the binding energies ranked in the order of Cu2+>Ni2+>Co2+>Fe2+>Cr2+>Zn2+>Mn2+; and for the same group of metal ions, the general trend of binding energies was Zn2+>Hg2+>Cd2+. Moreover, the amounts of charge transferred from S or N to transition metal cations are greater than that of O atoms. For Fe2+,Co2+,Ni2+,Cu2+,Zn2+,Cd2+ and Hg2+ complexes, the values of the Wiberg bond indices (WBIs) of M-S (M denotes metal cations) were larger than that of M-N and M-O; for Cr2+ complexes, most of the WBIs of M-O in complexes were higher than that of M-S and M-N. Furthermore, the changes in the electron configuration of the metal cations before and after chelate reaction revealed that Cu2+, Ni2+,Co2+ and Hg2+ had obvious tendencies to be reduced to Cu+,Ni+,Co+ and Hg+ during the coordination process. 相似文献
4.
Rishiram Ramanan Krishnamurthi Kannan Nadimuthu Vinayagamoorthy Kunga Mohan Ramkumar Saravana Devi Sivanesan Tapan Chakrabarti 《Biotechnology and Bioprocess Engineering》2009,14(1):32-37
Carbonic anhydrase enzyme, one of the fastest known enzymes, remains largely unexplored in prokaryotes when compared to its
mammalian counterparts despite its ubiquity. In this study, the enzyme has been purified from Bacillus subtilis SA3 using sequential Sephadex G-75 chromatography, DEAE cellulose chromatography, and sepharose-4B-L-tyrosinesulphanilamide
affinity chromatography and characterized to provide additional insights into its properties. The apparent molecular mass
of carbonic anhydrase obtained by SDS-PAGE was found to be approximately 37 kDa. Isoelectric focusing of the purified enzyme
revealed an isoelectric point (pI) of around 6.1 when compared with marker. The presence of metal ions such as Zn2+, Co2+, Cu2+, Fe3+, Mg2+, and anion SO4− increased enzyme activity while strong inhibition was observed in the presence of Hg2+, Cl−, HCO3−, and metal chelator EDTA. The optimum pH and temperature for the enzyme were found to be 8.3 and 37°C, respectively. Enzyme
kinetics with p-nitrophenyl acetate as substrate at pH 8.3 and 37°C determined the Vmax and Km values of the enzyme to be 714.28 μmol/mg protein/min and 9.09 mM, respectively. The Ki value for acetazolamide was 0.22 mM, compared to 0.099 mM for sulphanilamide. The results from N-terminal amino acid sequencing
imply the purified protein is a putative beta-carbonic anhydrase with close similarities to CAs from plants, microorganisms. 相似文献
5.
Chellappan S Jasmin C Basheer SM Kishore A Elyas KK Bhat SG Chandrasekaran M 《Journal of industrial microbiology & biotechnology》2011,38(6):743-752
An alkaline protease from marine Engyodontium album was characterized for its physicochemical properties towards evaluation of its suitability for potential industrial applications.
Molecular mass of the enzyme by matrix-assisted laser desorption ionization-mass spectrometry (MALDI-MS) analysis was calculated
as 28.6 kDa. Isoelectric focusing yielded pI of 3–4. Enzyme inhibition by phenylmethylsulfonyl fluoride (PMSF) and aprotinin
confirmed the serine protease nature of the enzyme. K
m, V
max, and K
cat of the enzyme were 4.727 × 10−2 mg/ml, 394.68 U, and 4.2175 × 10−2 s−1, respectively. Enzyme was noted to be active over a broad range of pH (6–12) and temperature (15–65°C), with maximum activity
at pH 11 and 60°C. CaCl2 (1 mM), starch (1%), and sucrose (1%) imparted thermal stability at 65°C. Hg2+, Cu2+, Fe3+, Zn2+, Cd+, and Al3+ inhibited enzyme activity, while 1 mM Co2+ enhanced enzyme activity. Reducing agents enhanced enzyme activity at lower concentrations. The enzyme showed considerable
storage stability, and retained its activity in the presence of hydrocarbons, natural oils, surfactants, and most of the organic
solvents tested. Results indicate that the marine protease holds potential for use in the detergent industry and for varied
applications. 相似文献
6.
Na'il Saleh 《Luminescence》2009,24(1):30-34
A water‐soluble, high‐output fluorescent sensor, based on a lumazine ligand with a thiophene substituent for Cd2+, Hg2+ and Ag+ metal ions, is reported. The sensor displays fluorescence enhancement upon Cd2+ binding (log β = 2.79 ± 0.08) and fluorescence quenching by chelating with Ag+ and Hg2+ (log β = 4.31 ± 0.15 and 5.42 ± 0.1, respectively). The mechanism of quenching is static and occurs by formation of a ground‐state non‐fluorescent complex followed by rapid intersystem crossing. The value of the Stern–Volmer quenching rate constant (kq) by Ag+ ions is close to 6.71 × 1012 mol/L/s at 298 K. The thermodynamic parameters (ΔG, ΔH and ΔS) were also evaluated and indicated that the complexation process is spontaneous, exothermic and entropically favourable. The quantitative linear relationship between the softness values of Klopman (σK) or Ahrland (σA) and the experimental binding constants (β) being in the order of Hg2+ > Ag+ > Cd2+ suggests that soft–soft interactions are the key for the observed sensitivity and selectivity in the presence of other metal ions, such as: Pb2+, Ni2+, Mn2+, Cu2+, Co2+, Zn2+ and Mg2+ ions. Copyright © 2008 John Wiley & Sons, Ltd. 相似文献
7.
Om Prakash Dinesh K. Vishwakarma 《Journal of plant biochemistry and biotechnology.》2001,10(2):147-149
Urease from the seeds of water melon was found to be inhibited by heavy metal ions like copper, lead, nickel and cobalt. The order of effectiveness of these metals as inhibitors was Cu2+ > Pb2+ > Ni2+ > Co2+. The inhibition by these ions was noncompetitive. Time — dependent interaction of urease with nickel and cobalt exhibited a biphasic inhibition behaviour in which approximately half of the initial activity was lost rapidly (within 2 min) and remainder in a slow phase. The inhibition was largely irreversible, hence could not be reversed by dialysis. These observations are suggestive of half-and-half distribution of — SH groups on the native enzyme resulting urease into asymmetric oligomeric molecule. 相似文献
8.
Elaine O’Connell Patrick Murray Charles Piggott Franck Hennequart Maria Tuohy 《Journal of applied phycology》2008,20(5):557-565
A β-N-acetylglucosaminidase produced by a novel fungal source, the moderately thermophilic aerobic ascomycete Talaromyces emersonii, was purified to apparent homogeneity. Submerged fermentation of T. emersonii, in liquid medium containing algal fucoidan as the main carbon source, yielded significant amounts of extracellular N-acetylglucosaminidase activity. The N-acetylglucosaminidase present in the culture-supernatant was purified by hydrophobic interaction chromatography and preparative
electrophoresis. The enzyme is a dimer with molecular weight and pI values of 140 and 3.85, respectively. Substrate specificity
studies confirmed the glycan specificity of the enzyme for N-acetylglucosamine. Michaelis-Menten kinetics were observed during enzyme-catalyzed hydrolysis of the fluorescent substrate
methylumbelliferyl-β-D-N-acetylglucosaminide at 50°C, pH 5.0 (Km value of 0.5 mM). The purified N-acetylglucosaminidase displayed activity over broad ranges of pH and temperature, yielding respective optimum values of pH
5.0 and 75°C. The T. emersonii enzyme was less susceptible to inhibition by N-acetylglucosamine and other related sugars than orthologs from other sources. The enzyme was sensitive to Hg2+, Co2+ and Fe3+. 相似文献
9.
Bacillus species producing a thermostable phytase was isolated from soil, boiled rice, and mezu (Korean traditinal koji). The activity of phytase increased markedly at the late stationary phase. An extracellular phytase from Bacillus sp. KHU-10 was purified to homogeneity by acetone precipitation and DEAE-Sepharose and phenyl-Sepharose column chromatographies. Its molecular weight was estimated to be 46 kDa on gel filtration and 44 kDa on SDS-polyacrylamide gel elctrophoresis. Its optimum pH and temperature for phytase activity were pH 6.5-8.5 and 40°C without 10 mM CaCl2 and pH 6.0-9.5 and 60°C with 10 mM CaCl2. About 50% of its original activity remained after incubation at 80°C or 10 min in the presence of 10 mM CaCl2. The enzyme activity was fairly stable from pH 6.5 to 10.0. The enzyme had an isoelectric point of 6.8. As for substrate specificity, it was very specific for sodium phytate and showed no activity on other phosphate esters. The K
m value for sodium phytate was 50 M. Its activity was inhibited by EDTA and metal ions such as Ba2+, Cd2+, Co2+, Cr3+, Cu2+, Hg2+, and Mn2+ ions. 相似文献
10.
Extracellular exoinulinase from Kluyveromyces marxianus YS-1, which hydrolyzes inulin into fructose, was immobilized on Duolite A568 after partial purification by ethanol precipitation
and gel exclusion chromatography on Sephadex G-100. Optimum temperature of immobilized enzyme was 55 °C, which was 5 °C higher
than the free enzyme and optimal pH was 5.5. Immobilized biocatalyst retained more than 90% of its original activity after
incubation at 60 °C for 3 h, whereas in free form its activity was reduced to 10% under same conditions, showing a significant
improvement in the thermal stability of the biocatalyst after immobilization. Apparent K
m values for inulin, raffinose and sucrose were found to be 3.75, 28.5 and 30.7 mM, respectively. Activation energy (E
a) of the immobilized biocatalyst was found to be 46.8 kJ/mol. Metal ions like Co2+ and Mn2+ enhanced the activity, whereas Hg2+ and Ag2+ were found to be potent inhibitors even at lower concentrations of 1 mM. Immobilized biocatalyst was effectively used in
batch preparation of high fructose syrup from Asparagus racemosus raw inulin and pure inulin, which yielded 39.2 and 40.2 g/L of fructose in 4 h; it was 85.5 and 92.6% of total reducing sugars
produced, respectively. 相似文献
11.
《Biochimica et Biophysica Acta (BBA) - Protein Structure and Molecular Enzymology》1996,1292(1):188-196
We studied the transition metal ion requirements for activity and sulfhydryl group reactivity in phosphoenolpyruvate carboxykinase (PEP-carboxykinase; ATP:oxaloacetate carboxylase (transphosphorylating), EC 4.1.1.49), a key enzyme in the energy metabolism of the protozoan parasite Trypanosoma (Schizotrypanum) cruzi. As for other PEP-carboxykinases this enzyme has a strict requirement of transition metal ions for activity, even in the presence of excess Mg2+ ions for the carboxylation reaction; the order of effectiveness of these ions as enzyme activators was: Co2+ > Mn2+ > Cdu2+ > Ni2+ ⪢ Fe2+ > VO2+, while Zn2+ and Ca2+ had no activating effects. When we investigated the effect of varying the type or concentration of the transition metal ions on the kinetic parameters of the enzyme the results suggested that the stimulatory effects of the transition metal center were mostly associated with the activation of the relatively inert CO2 substrate. The inhibitory effects of 3-mercaptopicolinic acid (3MP) on the enzyme were found to depend on the transition metal ion activator: for the Mn2+ activated enzyme the inhibition was purely non-competitive (Kii = Kis) towards all substrates, while for the Co2+-activated enzyme the inhibitor was much less effective, produced a mixed-type inhibition and affected differentially the interaction of the enzyme with its substrates. The modification of a single, highly reactive, cysteine per enzyme molecule by 5,5′-dithiobis(2-nitro-benzoate) (DTNB) lead to an almost complete inhibition of Mn2+-activated T. cruzi PEP-carboxykinase; however, in contrast with the results of previous studies in vertebrate and yeast enzymes, the substrate ADP slowed the chemical modification and enzyme inactivation but did not prevent it. PEP and HCO3− had no significant effect on the rate or extent of the enzyme inactivation. The kinetics of the enzyme inactivation by DTNB was also dependent on the transition metal activator, being much slower for the Co2+-activated enzyme than for its Mn2+-activated counterpart. When the bulkier but more hydrophobic reagent N-(7-dimethylamino-4-methylcoumarinyl)maleimide (DACM) was used the enzyme was slowly and incompletely inactivated in the presence of Mn2+ and ADP afforded almost complete protection from inactivation; in the presence of Co2+ the enzyme was completely resistant to inactivation. Taken together, our results indicate that the parasite enzyme has a specific requirement of transition metal ions for activity and that they modulate the reactivity of a single, essential thiol group, different from the hyperreactive cysteines present in vertebrate or yeast enzymes. 相似文献
12.
Nguyen VN Oh IJ Kim YJ Kim KY Kim YC Park RD 《Journal of industrial microbiology & biotechnology》2009,36(2):195-203
Two extracellular chitinases were purified from Paecilomyces variotii DG-3, a chitinase producer and a nematode egg-parasitic fungus, to homogeneity by DEAE Sephadex A-50 and Sephadex G-100 chromatography.
The purified enzymes were a monomer with an apparent molecular mass of 32 kDa (Chi32) and 46 kDa (Chi46), respectively, and
showed chitinase activity bands with 0.01% glycol chitin as a substrate after SDS-PAGE. The first 20 and 15 N-terminal amino
acid sequences of Chi32 and Chi46 were determined to be Asp-Pro-Typ-Gln-Thr-Asn-Val-Val-Tyr-Thr-Gly-Gln-Asp-Phe-Val-Ser-Pro-Asp-Leu-Phe
and Asp-Ala-X-X-Tyr-Arg-Ser-Val-Ala-Tyr-Phe-Val-Asn-Trp-Ala, respectively. Optimal temperature and pH of the Chi32 and Chi46
were found to be both 60°C, and 2.5 and 3.0, respectively. Chi32 was almost inhibited by metal ions Ag+ and Hg2+ while Chi46 by Hg2+ and Pb2+ at a 10 mM concentration but both enzymes were enhanced by 1 mM concentration of Co2+. On analyzing the hydrolyzates of chitin oligomers [(GlcNAc)
n
, n = 2–6)], it was considered that Chi32 degraded chitin oligomers as an exo-type chitinase while Chi46 as an endo-type chitinase. 相似文献
13.
An investigation on the properties of an alkaline protease secreted by Bacillus circulans BM15 strain isolated from a mangrove sediment sample was carried out in order to characterize the enzyme and to test its
potency as a detergent additive. The protease was purified to apparent homogeneity by ammonium sulphate precipitation and
was a 30-kDa protease as shown by SDS-PAGE and its proteolytic activity was detected by casein zymography. It had optimum
activity at pH 7, was stable at alkaline pH range (7 to 11), had optimum temperature of activity 40°C and was stable up to
a temperature of 55°C after incubation for one hour. Hg2+, Zn2+, Co2+, and Cu2+completely inhibited the enzyme activity, while Ca2+, Mg2+, K+ and Fe3+ were enhancing the same. The serine protease inhibitor PMSF and metal chelator EDTA inhibited the activity of this protease
while the classic metalloprotease inhibitor 1, 10 phenanthroline did not show inhibition. The enzyme was stable in SDS, Triton-X-100
and H2 O2 as well as in various commercial detergents after incubation for one hour. The extracellular production of the enzyme, the
pH and temperature stability and stability in presence of oxidants, surfactants and commercial detergents suggest its possible
use as a detergent additive. 相似文献
14.
Eliane Bahbouth Brigitte Siwek Marie-Claire De Pauw-Gillet Enrico Sabbioni Roger Bassleer 《Biological trace element research》1993,36(2):191-201
The effects of fourteen metal ions (As3+, As5+, Cd2+, Co2+, Cr3+, Cr6+, Hg2+, Li+, Mg2+, Mn2+, Ni2+, Se4+, V5+, VO2+) on the proliferation and differentiation in mouse B16 melanoma cells cultivated in vitro were analyzed. Cell number assays,
melanin, and protein measurements, a 3(4,5-dimethylthiazol-2-yl)-2,5 diphenyltetrazolium bromide reduction test (MTT survival
test), and a clonal growth assay were performed. At 10−4
M, metal ions such as As3+, As5+, Cd2+, Cr6+, Se4+, V5+, VO2+, and, to a minor extent, Li+, Hg2+, and Co2+ significantly reduced the number of the B16 melanoma cells. For the same molar concentration, the order of the levels of
cell toxicity of the metal compounds to B16 cells as measured by the MTT test was as follows: Hg2+>Cr6+=Cd2+>As3+, As5+>V5+, VO2+>Se4+=Ni2+=Co2+=Li+. An increased synthesis of melanin in B16 cells was noted after incubation with Co2+, Ni2+, Cd2+, and Li+, whereas Se4+ had, on the contrary, an inhibiting effect on melanogenesis. 相似文献
15.
Jeyagowri Kiddinamoorthy Alfredo J. Anceno Gulelat D. Haki Sudip K. Rakshit 《World journal of microbiology & biotechnology》2008,24(5):605-612
Production of extracellular xylanase from Bacillus sp. GRE7 using a bench-top bioreactor and solid-state fermentation (SSF) was attempted. SSF using wheat bran as substrate
and submerged cultivation using oat-spelt xylan as substrate resulted in an enzyme productivity of 3,950 IU g−1 bran and 180 IU ml−1, respectively. The purified enzyme had an apparent molecular weight of 42 kDa and showed optimum activity at 70°C and pH
7. The enzyme was stable at 60–80°C at pH 7 and pH 5–11 at 37°C. Metal ions Mn2+ and Co2+ increased activity by twofold, while Cu2+ and Fe2+ reduced activity by fivefold as compared to the control. At 60°C and pH 6, the K
m for oat-spelt xylan was 2.23 mg ml−1 and V
max was 296.8 IU mg−1 protein. In the enzymatic prebleaching of eucalyptus Kraft pulp, the release of chromophores, formation of reducing sugars
and brightness was higher while the Kappa number was lower than the control with increased enzyme dosage at 30% reduction
of the original chlorine dioxide usage. The thermostability, alkali-tolerance, negligible presence of cellulolytic activity,
ability to improve brightness and capacity to reduce chlorine dioxide usage demonstrates the high potential of the enzyme
for application in the biobleaching of Kraft pulp. 相似文献
16.
Bacterial allantoinase (ALLase; EC 3.5.2.5), which catalyzes the conversion of allantoin into allantoate, possesses a binuclear
metal center in which two metal ions are bridged by a posttranslationally carboxylated lysine. Here, we characterized ALLase
from Escherichia coli BL21. Purified recombinant ALLase exhibited no activity but could be activated when preincubating with some metal ions before
analyzing its activity, and was in the order: Mn2+- ≫ Co2+- > Zn2+- > Ni2+- > Cd2+- ~Mg2+-activated enzyme; however, activity of ALLase (Mn2+-activated form) was also significantly inhibited with 5 mM Co2+, Zn2+, and Cd2+ ions. Activity of Mn2+-activated ALLase was increased by adding the reducing agent dithiothreitol (DTT), but was decreased by treating with the
sulfhydryl modifying reagent N-ethylmaleimide (NEM). Inhibition of Mn2+-activated ALLase by chelator 8-hydroxy-5-quinolinesulfonic acid (8-HQSA), but not EDTA, was pH-dependent. Analysis of purified
ALLase by gel filtration chromatography revealed a mixture of monomers, dimers, and tetramers. Substituting the putative metal
binding residues His59, His61, Lys146, His186, His242, and Asp315 with Ala completely abolished the activity of ALLase, even
preincubating with Mn2+ ions. On the basis of these results, as well as the pH-activity profile, the reaction mechanism of ALLase is discussed and
compared with those of other cyclic amidohydrolases. 相似文献
17.
Xia Hu Chunyan Wang Le Wang Ranran Zhang Hui Chen 《World journal of microbiology & biotechnology》2014,30(4):1285-1290
The bark beetle Dendroctonus armandi is able to kill living Pinus armandi and has caused serious damage to pine forest in Northern China. As the most important symbiotic fungus of D. armandi, Leptographium qinlingensis plays an important role in the invasion process of the bark beetle. The laccase secreted by it are involved in lignin degradation to provide utilizable nutrition for D. armandi, and catalyze some biochemical reactions, causing the damages of tree tissue. In present study, the extracellular laccase of L. qinlingensis was purified by using the ammonium sulfate precipitation and DEAE-cellulose (DE-52) column chromatography. Furthermore, the effects of temperature, pH value and metal ions on it were investigated and characterized. The purified enzyme exerted its optimal activity with guaiacol. The catalytic efficiencies Km and Vmax determined for substrate guaiacol were 15.4 μM and 372.9 IU mg?1, respectively. The optimum pH and temperature for the purified enzyme was 4.4 and 45 °C, respectively, with the highest enzyme specific activity of 7,000 IU mg?1. Moreover, the metal ions, Co2+, Mn2+, Ca2+, Mg2+, Fe2+ and Cd2+, especially Hg2+, showed significantly inhibition effects on its activity. To understand the characteristics of this laccase might provide an opportunity and theoretical basis to promote integrated pest management of D. armandi. 相似文献
18.
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
19.
A 43 kDa α-amylase was purified from Tinospora cordifolia by glycogen precipitation, ammonium sulfate precipitation, gel filtration chromatography, and HPGPLC. The enzyme was optimally active in pH 6.0 at 60 °C and had specific activity of 546.2 U/mg of protein. Activity was stable in the pH range of 4-7 and at temperatures up to 60 °C. PCMB, iodoacetic acid, iodoacetamide, DTNB, and heavy metal ions Hg2+ > Ag+ > Cd2+ inhibited enzyme activity while Ca2+ improved both activity and thermostability. The enzyme was a thiol amylase (3 SH group/mole) and DTNB inhibition of activity was released by cysteine. N-terminal sequence of the enzyme had poor similarity (12-24%) with those of plant and microbial amylases. The enzyme was equally active on soluble starch and amylopectin and released maltose as the major end product. 相似文献
20.
The ability of trout to metabolize aniline in vitro in the presence of some divalent metal ions was investigated in the liver microsomes of rainbow trout, Salmo gairdneri. Trout liver microsomes were highly capable of catalyzing aniline hydroxylation to p-aminophenol with a specific activity of 0.068 nmoles/min per mg of microsomal protein in potassium phosphate buffer, pH 7.4 at 25°C. The activity of the aniline hydroxylase system was competitively inhibited by Hg+2, Ni+2, Cd+2, and Zn+2, while Cu+2 and Fe+3 seemed to inhibit the activity noncompetitively at 1 mM aniline concentrations. IC50 values at fixed aniline concentration were estimated to be 0.45 mM for Hg+2, Ni+2, and Cd+2, 1.8 mM for Zn+2 and Fe+3, and 1.3 mM for Cu+2. Eadie-Hofstee plots gave identical Vmax values of approximately 0.046 nmol/min per mg of protein while Km values were increased in the presence of Hg+2, Ni+2, CD+2, and Zn+2, indicating competitive inhibition. Both Km and Vmax values were affected by Fe+3 and Cu+2, suggesting noncompetitive inhibition. Ki values extracted from the Dixon plots were determined t be 0.23, 0.43, and 0.65 mM for Hg+2, Ni+2, and Cd+2, respectively, providing the most effective inhibition on the aniline hydroxylase system among studied metal ions. The Ki values were much higher in the presence of others. The results indicate a selective inhibition of the aniline hydroxylase system of trout liver microsomes by divalent metal ions. © 1997 John Wiley & Sons, Inc. 相似文献