共查询到20条相似文献,搜索用时 31 毫秒
1.
Bush LP 《Plant physiology》1969,44(3):347-350
Succinyl CoA synthetase from Nicotiana tabacum exhibited a requirement for univalent and divalent cations. Mn2+ replaced Mg2+ in the assay medium and Co2+ and Ca2+ partially replaced Mg2+. Addition of Zn2+ resulted in no enzyme activity. The enzyme was activated by univalent cations K+, Rb+, NH4+, and Na+; Li+ showed little or no activation. Maximum enzyme activity varied significantly with potassium salts of different anions. Greatest activation was obtained with K3PO4 and, respectively, KCl, KNO3, K2SO4 and KF exhibited steadily decreasing enzyme activation. 相似文献
2.
The changes of granule bound starch synthetase activity in cucumber leaves (Cucumis sativus L. cv. Suisei No. 2) were investigated during ammonium toxicity. Generally speaking the quantity of starch granules of injured plants were less than that of normal plants. ADPG is a more effective glucose donor to starch synthesis than UDPG. It was found that the starch synthetase activity of injured plants was decreased compared to the normal plants. This variation of enzyme activity was higher when UDPG was used as glucose donor. The addition of K+ and NH4+ generally inhibited the enzyme activity when UDPG was used as glucose donor, but stimulated it when ADPG was used. This stimulation was found to be more effective in enzymes prepared from injured plants than from normal plants. The level of potassium bound to starch granules was not changed markedly between normal and injured plants. 相似文献
3.
Solubilization and partial purification of yeast chitin synthetase. Confirmation of the zymogenic nature of the enzyme. 总被引:13,自引:0,他引:13
Chitin synthetase was solubilized with digitonin from a particulate yeast fraction. The solubilized enzyme, which did not sediment at 200,000 X g and emerged after the void volume in a Sepharose 6B column, was active only after treatment with a protease. This confirms that chitin synthetase exists in the plasma membrane as a zymogen and that initiation of the chitin septum occurs by localized activation of the enzyme. By differential extraction with sodium cholate and digitonin, followed by chromatography on Sepharose 6B, a 20-fold purification of the enzyme was achieved with respect to the crude particles. The purified enzyme showed a requirement for a phospholipid; phosphatidylserine and lysophosphatidylserine were the best activators. Unsaturated fatty acids strongly inhibited synthetase activity, whereas their saturated counterparts were inert. The solubilized enzyme catalyzed the formation of insoluble chitin in the absence of added primer. The synthetic polysaccharide was examined by electron microscopy and found to consist of lozenge-shaped particles about 60 nm long and 10 nm wide. 相似文献
4.
Electron photomicrographs of endosperm tissue from germinating seed of Ricinus communis L. cv. Hale show proplastids which contain prominent starch grains. The content of starch in endosperm tissue increased from 500 micrograms per seed, in imbibed seed, to 1,100 micrograms per seed in 5-day-old seedlings. The maximum net rate of starch deposition was 1.1 nanomoles glucose incorporated per minute per seed. About 200 micrograms of starch remained in the endosperm 9 days after imbibition. Starch content followed the same developmental pattern as the content of sucrose, free reducing sugars, and other metabolic processes found in this tissue. Two key enzymes of starch synthesis, adenosine diphosphoglucose (ADPG) pyrophosphorylase and ADPG-starch glucosyl transferase (starch synthetase) exhibited maximum activities at 4 and 5 days after germination, respectively. The maximum activity of ADPG pyrophosphorylase was 8.17 nanomoles ADPG formed per minute per seed, whereas starch synthetase exhibited an activity of 125 nanomoles glucose incorporated per minute per seed. These levels of enzyme activity are sufficient to account for the starch synthesis observed. Other enzymes which may be involved in starch synthesis include 3-phosphoglycerate kinase which showed an activity of 8.76 units per seed, triose-P isomerase (2.56 units per seed), fructose-1,6-bisphosphate aldolase (0.99 units per seed), fructose-1,6-bisphosphatase (0.23 units per seed), phosphoglucose isomerase (12.6 units per seed), and phosphoglucomutase (9.72 units per seed). The activities of these enzymes were similar to previously reported values.
Starch synthetase was found in association with the fraction containing proplastids isolated from endosperm tissue. Of the total starch synthetase activity in the endosperm, 38% was particulate. Forty-four% of the total particulate activity of starch synthetase placed on sucrose gradients was associated with the band containing proplastids. The proplastids contained 98% of the ribulose 1,5-bisphosphate carboxylase carboxylase activity placed on the gradient.
相似文献5.
Chicken liver pyruvate carboxylase has an absolute requirement for short-chain acyl coenzyme A (CoA), whereas the same enzyme from yeast has less stringent requirements. The yeast enzyme has now been studied in an effort to elucidate the mechanism by which acyl-CoA stimulates pyruvate carboxylase activity. Yeast pyruvate carboxylase has an apparent basal level of activity above which CoA and acyl-CoAs of 2-20 carbons activate; the concentration of acyl-CoA required for half-maximum activation (K0.5) decreases as the chain length of the acyl moiety increases to 16 carbons. Activation of yeast pyruvate carboxylase by acyl-CoA is brought about in part by increasing the affinity of pyruvate carboxylase for two substrates, bicarbonate and pyruvate. The affinity of pyruvate carboxylase for bicarbonate is also increased by potassium ions. The observation of only low levels of activity in the absence of acyl-CoA or potassium ion leads to the conclusion that the basal activity so frequently referred to is probably due to the presence of activating monovalent cations. Pyruvate carboxylase from yeast probably has an absolute requirement for monovalent cations or acyl-CoA with a combination of the two being required for optimum conditions for maximal activity. Stimulation by acyl-CoA and inhibition by aspartate are mutually antagonistic with each affecting the activation or inhibition constant and the degree of cooperativity brought about by the other. The enzyme from liver is unaffected by aspartate. 相似文献
6.
ADPglucose: α-1,4-glucan α-4-glucosyltransferases (starch synthetases) from leaves of Vitis vinifera and leaves and kernels of Zea mays were chromatographed on DEAE-cellulose columns. One form of the enzyme was present in grape leaves having activity both in the presence and absence of primer. Two forms were present in both leaves and kernels of maize. The second peak of activity in maize leaves and the first peak in maize kernels synthesized a polyglucan in the absence of primer. A peak of branching enzyme (Q-enzyme) occurred between the two starch synthetase peaks with both tissues. When fractions containing starch synthetase and branching enzyme were added to the first leaf starch synthetase peak, up to 100-fold activation of the unprimed reaction occurred. Branching enzyme did not stimulate the unprimed activity of the first kernel peak and no branching enzyme could be detected in this peak. The unprimed product was a branched polyglucan with mainly α-1,4-links. 相似文献
7.
An investigation has been made to determine the effectiveness of univalent cations as cofactors for the inductive synthesis of nitrate reductase. In these experiments K(+) functions more effectively as the univalent cation activator than other univalent cations. Substitution of Rb(+) for K(+) resulted in enzyme formation at a rate of about one-half of that obtained with K(+). Sodium, Li(+), or NH(4) (+) either failed to stimulate or completely inhibited the inductive formation of the enzyme. When no univalent cations were present in the induction medium, enzyme formation was delayed for an initial 3-hour period in contrast to the normal one-hour delay in enzyme formation where adequate K(+) was present in the induction medium.During the period of inductive formation of nitrate reductase the activity of pyruvic kinase, a constitutive enzyme, was assayed under conditions where adequate K(+) was present. Results indicate that the presence of the different univalent cations in the induction medium had no striking effect on the activity of this enzyme during the induction period. 相似文献
8.
Selective activation of particulate guanylate cyclase by a specific class of porphyrins 总被引:1,自引:0,他引:1
S A Waldman M S Sinacore J A Lewicki L Y Chang F Murad 《The Journal of biological chemistry》1984,259(7):4038-4042
Guanylate cyclase was activated 3- to 10-fold by hemin in a dose-dependent manner in membranes prepared from homogenates of rat lung, C6 rat glioma cells, or B103 rat neuroblastoma cells. Maximum activation was observed with 50 to 100 microM hemin with higher concentrations being inhibitory. Activation was observed when Mg2+-GTP but not when Mn2+-GTP was used as the substrate. Increased enzyme activity reflected selective activation of the particulate form of guanylate cyclase; hemin inhibited the soluble form of guanylate cyclase 70 to 90% over a wide range of concentrations. Activation was not secondary to proteolysis since a variety of protease inhibitors failed to alter stimulation by hemin. Protophorphyrin IX had little effect on particulate guanylate cyclase activity and sodium borohydride almost completely abolished hemin-dependent activation. These data suggest a requirement for the ferric form of the porphyrin-metal chelate for activation. However, agents which interact with the iron nucleus of porphyrins, such as cyanide, had little effect on the ability of hemin to activate guanylate cyclase. The stimulatory effects of hemin were observed in the presence of detergents such as Lubrol-PX, and highly purified particulate enzyme could be activated to the same extent as enzyme in native membranes. These data suggest that the interaction of porphyrins with particulate guanylate cyclase is complex in nature and different from that with the soluble enzyme. 相似文献
9.
P F Duggan 《The Journal of biological chemistry》1977,252(5):1620-1627
The effects of monovalent cations on calcium uptake by fragmented sarcoplasmic reticulum have been clarified. Homogenization of muscle tissue in salt-containing solutions leads to contamination of this subcellular fraction with actomyosin and mitochondrial membranes. When, in addition, inorganic cations are contributed by the microsomal suspension and in association with nucleotide triphosphate substrates there is an apparent inhibition of the calcium transport system by potassium and other cations. However, when purified preparations were obtained after homogenization in sucrose medium followed by centrifugation on a sucrose density gradient in a zonal rotor, calcium uptake and the associated adenosine triphosphatase activity were considerably activated by potassium and other univalent cations. When plotted against the log of the free calcium concentration there was only a slight increase in calcium uptake and ATPase activity in the absence of potassium ions but sigmoid-shaped curves were obtained in 100 mM K+ with half-maximal stimulation occurring at 2 muM Ca2+ for both calcium uptake and ATPase activity. The augmentation in calcium uptake was not due to an ionic strength effect as Tris cation at pH 6.6 was shown to be inactive in this respect. Other monovalent cations were effective in the order K+ greater than Na+ greater than NH4+=Rb+=Cs+ greater than Li+ with half-maximal stimulation in 11 mM K+, 16 mM Na+, 25 mM NH4+, Rb+, and Cs+ and in 50 mM Li+. There was nos synergistic action between K+ AND Na+ ions and both calcium uptak and associated ATPase were insensitive to ouabain. Thallous ions stimulate many K+-requiring enzymes and at one-tenth the concentration were nearly as effective as K+ ions in promoting calcium uptake. The ratio of Ca2+ ions transported to P1 released remained unchanged at 2 after addition of K+ ions indicating an effect on the rate of calcium uptake rather than an increased efficiency of uptake. In support of this it was found that during the stimulation of calcium uptake by Na+ ions there was a reduction in the steady state concentration of phosphorylated intermediate formed from [gamma-32P]ATP. It is considered that there is a physiological requirement for potassium ions in the relaxation process. 相似文献
10.
The ion-dependent enzymes, activities of which are affected by ions, were entrapped in the inside aqueous phase of reverse-phase evaporation vesicles (REVs) to protect the enzyme activities from inhibitory cations. Their activities were still preserved in the presence of the inhibitory cations because the permeabilities of the inhibitory cations through the lipid membranes of REVs were much lower than those of substrates and products. The REVs were relatively stable in hypertonic condition, while they were unstable in hypotonic condition. Changes in osmotic pressure difference largely affected solute permeabilities of REVs. The shrinking and swelling of REVs resulting from osmotic pressure differences led to these changes in stabilities and solute permeabilities. Although the entrapment of the enzyme into REVs showed good protective effects against divalent cations, it was not effective against univalent cations. The measurements of cation permeabilities revealed that the enzyme trapped in the bilayer regions of REVs acts as a selective channel for the univalent cations. The REVs containing the enzyme could be used without any activity losses in a hollow fiber module. 相似文献
11.
以"凤丹"牡丹根际土壤中分离筛选到的产脂肪酶菌株Pseudomonas sp. RYXP作为出发菌株,对其进行了紫外线诱变选育,并采用单因素试验和正交试验方法对活性最强正突变株的产脂肪酶基本特性进行了测定。结果表明,出发菌株Pseudomonas sp. RYXP的紫外线诱变最佳条件为:15 W紫外灯30 cm距离照射1 min;将产脂肪酶活性最强的正突变菌株编号为RYXP-3,单因素试验表明RYXP-3产脂肪酶适宜的碳源为玉米淀粉,适宜氮源为豆饼粉,适宜的磷酸二氢钾含量是0.3%,适宜的初始pH值为7;正交试验表明RYXP-3的最佳的产酶培养基成分组成是:玉米淀粉7%,豆饼粉3%,磷酸二氢钾0.3%,初始pH值为8。在优化方案A1B2C2D3产酶条件下,突变株RYXP-3最高的产酶活性达到56.1 U/mL。突变菌株RYXP-3可作为产油牡丹"凤丹"专用促生菌肥开发的备选资源菌株。 相似文献
12.
A glutathione-S-transferase involved in atrazine conjugation was purified 43-fold from corn with a total yield of 36%. The purified enzyme has a MW of 45 000 as determined by gel filtration. The estimated activation energy of the enzyme is 6.4 kcal/mol and the optimum pH for activity between 8 and 8.5. Substrate specificity studies with s-triazines indicated that atrazine was the best substrate followed by simazine and propazine. The Cl group at the 2-position was essential for enzyme activity, and replacement by a SCH3 group resulted in a total loss of activity. The absence of an alkyl group resulted in a reduction of conjugation and 2-chloro-4,6-bis-amino-s-triazine was the poorest substrate. With insecticidal substrates (organophosphates), conjugating activity was observed only with diazinon and little or no activity was observed with ethyl parathion, malathion and etrimfos. No activity was found using methyl iodide as a substrate. The purified enzyme has properties similar to those of an aryl-S-transferase. Quinones were inhibitors of this enzyme. 相似文献
13.
Aspergillus sp GP-21 produced a raw-starch digesting amyloglucosidase which showed optimum activity at 65°C and pH 5.0–5.5. At 50°C the
enzyme converted about 40% of raw corn starch to glucose within 48 h. Enzyme production was studied in solid state fermentation
using wheat bran. Productivity was affected by the level of moisture, incubation temperature and the presence or absence of
supplements. Maximum enzyme production was observed at a moisture level of 75% and at 30°C. Enzyme production was stimulated
by supplementing wheat bran with 0.25% proteose peptone, 1% trace mineral solution, 0.01% CaCl2 and 0.01% MgSO4.
Received 27 October 1998/ Accepted in revised form 15 March 1999 相似文献
14.
De Sousa MO Santoro MM De Souza Figueiredo AF 《Journal of enzyme inhibition and medicinal chemistry》2004,19(4):317-325
Hydrolysis of D-valyl-L-leucyl-L-arginine p-nitroanilide by human tissue kallikrein (hK1) was studied in the absence and in the presence of increasing concentrations of the following chloride salts: sodium, potassium, calcium, magnesium and aluminium. The data indicate that the inhibition of hK1 by sodium, potassium, calcium and magnesium is linear competitive and that divalent cations are more potent inhibitors of hK1 than univalent cations. However the inhibition of hK1 by aluminium cation is linear mixed, with the cation being able to bind to both the free enzyme and the ES complex. This cation was the best hK1 inhibitor. Aluminium is not a physiological cation, but is a known neurotoxicant for animals and humans. The neurotoxic actions of aluminium may relate to neuro-degenerative diseases. 相似文献
15.
海洋环境来源的淀粉酶AmyP对生玉米 淀粉的降解特性 总被引:1,自引:0,他引:1
来自海洋宏基因组文库的 α-淀粉酶(AmyP)属于最新建立的糖苷水解酶亚家族GH1337。AmyP 是一个生淀粉降解酶,能有效降解玉米生淀粉。在最适反应条件 pH 7.5和 40 °C 下,生玉米淀粉的比活达到 39.6 ± 1.4 U/mg。酶解反应动力学显示 AmyP 可以非常快速的降解生玉米淀粉。对 1%的生玉米淀粉仅需要 30 min;4%和 8%的生玉米淀粉只需 3 h。DTT 可以显著提高 AmyP 对生玉米淀粉的降解活性,1% DTT 促使活性增加 1倍。根据电镜观察和产物分析,认为 AmyP 是以内腐蚀的模式降解生玉米淀粉颗粒,释放出葡萄糖、麦芽糖和麦芽三糖作为终产物。 相似文献
16.
1. A purification procedure for DNA nucleotidyltransferase from Landschütz ascites-tumour cells is described. The enzyme can be separated from endogenous nucleic acid and from triphosphatase and deoxyribonuclease activities measurable at pH7.5. 2. The basic properties of the nucleotidyltransferase reaction are as follows. The enzyme has optimum activity at pH7.2-7.4. It displays an absolute requirement for DNA-primer, thermally-denatured DNA serving three to ten times as efficiently in this respect as native DNA. Maximum synthesis of polydeoxyribonucleotide occurs in the presence of all four deoxyribonucleoside 5'-triphosphates, but a limited incorporation of mononucleotide into polynucleotide is observed when the system is provided with only one triphosphate, or with various combinations of mono-, di- and tri-phosphates. The reaction requires the presence of a bivalent cation, and of those tested, Mg(2+) ions were by far the most effective. Manganous ions promoted synthesis but to a much smaller extent. Calcium ions did not support synthesis at all. At the appropriate concentrations, the univalent cations (sodium and potassium) stimulated the reaction by 25% and 125% respectively. The presence of EDTA in the reaction mixture stimulates the system five- to ten-fold. 3. The storage characteristics of the enzyme (as well as the activities of the various fractions) improve markedly if EDTA and 2-mercaptoethanol are included in the enzyme solution and in all preparative buffer solutions. 4. The enzyme loses more than 95% of its activity after heating for 1min. at 45 degrees . If the heating is conducted in the presence of DNA, the enzyme becomes relatively heat-resistant (presumably as a consequence of complex-formation with the DNA) and may actually display an activation effect. This is discussed in relation to a possible molecular conformation of the enzyme. 5. The product of the nucleotidyltransferase reaction is precipitable by acid or ethanol, and is susceptible to the actions of deoxyribonucleases I and II, snake-venom and spleen phosphodiesterases, and micrococcal nuclease. It forms a band in a density gradient of caesium chloride at a density similar to that of the DNA-primer. 6. By the criteria of nearest-neighbour frequency analyses, the product of the nucleotidyltransferase reaction has the characteristics to be expected of a polynucleotide synthesized in accordance with the template directions of the primer. 相似文献
17.
C P Selitrennikoff 《Biochimica et biophysica acta》1979,571(2):224-232
Chitin synthase (UDP-2-acetamido-2-deoxy-D-glucose:chitin 4-beta-acetamidodeoxy-D-glucosyltransferase, EC 2.4.1.16) activity from the wall-less variant of Neurospora crassa (slime) was partially characterized. The slime enzyme activity was found to be similar to that reported for slime-like and wild-type chitin synthase activities with respect to the following: specific activity, particulate cell-fraction localization, activation by N-acetylglucosamine, apparent Km with respect to substrate, pH optimum and ion requirement. It appears that the phenotype of slime cannot be solely accounted for by the absence of chitin synthase enzyme activity. 相似文献
18.
Marinez de Oliveira Sousa Marcelo Matos Santoro 《Journal of enzyme inhibition and medicinal chemistry》2013,28(4):317-325
Hydrolysis of D-valyl-L-leucyl-L-arginine p-nitroanilide by human tissue kallikrein (hK1) was studied in the absence and in the presence of increasing concentrations of the following chloride salts: sodium, potassium, calcium, magnesium and aluminium. The data indicate that the inhibition of hK1 by sodium, potassium, calcium and magnesium is linear competitive and that divalent cations are more potent inhibitors of hK1 than univalent cations. However the inhibition of hK1 by aluminium cation is linear mixed, with the cation being able to bind to both the free enzyme and the ES complex. This cation was the best hK1 inhibitor. Aluminium is not a physiological cation, but is a known neurotoxicant for animals and humans. The neurotoxic actions of aluminium may relate to neuro-degenerative diseases. 相似文献
19.
λ-Glutamylcysteine synthetase in higher plants: catalytic properties and subcellular localization 总被引:1,自引:0,他引:1
λ-Glutamylcysteine synthetase activity (EC 6.3.2.2) was analysed in Sephacryl S-200 eluents of extracts from cell suspension
cultures ofNicotiana tabacum L. cv. Samsun by determination of λ-glutamylcysteine as its monobromobimane derivative. The enzyme has a relative molecular
mass (Mr) of 60000 and exhibits maximal activity at pH 8 (50% at pH 7.0 and pH9.0) and an absolute requirement for Mg2+. With 0.2mM Cd2+ or Zn2+, enzyme activity was reduced by 35% and 19%, respectively. Treatment with 5 mM dithioerythritol led to a heavy loss of activity
and to dissociation into subunits (Mr 34000). Buthionine sulfoximine andl-methionine-sulfoximine, known as potent inhibitors of λ-glutamylcysteine synthetase from mammalian cells, were found to be
effective inhibitors of the plant enzyme too. The apparent Km values forl-glutamate,l-cysteine, and α-aminobutyrate were, respectively, 10.4mM, 0.19 mM, and 6.36 mM. The enzyme was completely inhibited by glutathione
(Ki=0.42 mM). The data indicate that the rate of glutathione synthesis in vivo may be influenced substantially by the concentration
of cysteine and glutamate and may be further regulated by feedback inhibition of λ-glutamylcysteine synthetase by glutathione
itself. λ-Glutamylcysteine synthetase is, like glutathione synthetase, localized in chloroplasts as well as in the cytoplasm.
Chloroplasts fromPisum sativum L. isolated on a Percoll gradient contained about 72% of the λ-glutamylcysteine synthetase activity in leaf cells and 48%
of the total glutathione synthetase activity. In chloroplasts ofSpinacia oleracea L. about 61% of the total λ-glutamylcysteine synthetase activity of the cells were found and 58% of the total glutathione
synthetase activity. These results indicate that glutathione synthesis can take place in at least two compartments of the
plant cell.
Dedicated to Professor A. Prison on the occasion of his 80th birthday 相似文献
20.
A reconstitution system for activation of the O2(.-)-generating oxidase from bovine polymorphonuclear neutrophils (PMN) is described. This system consisted of three components, namely, a particulate fraction enriched in plasma membrane, a supernatant fluid (cytosolic fraction) recovered by high-speed centrifugation from sonicated resting bovine PMN, and arachidonic acid. The pH optimum (7.8) and the Km value for NADPH (45 microM) of the activated oxidase were virtually the same as those found in the purified enzyme. All three components had to be present during the preincubation for elicitation of oxidase activity. A further enhancement of oxidase activity was observed with the addition of nonhydrolyzable GTP analogues, such as guanosine 5'-O-(3-thiotriphosphate) (GTP-gamma-S) and guanosine 5'-(beta, gamma-imidotriphosphate) (GMP-PNP), to the preincubation medium. In contrast, GDP-beta-S drastically decreased oxidase activation. In a two-stage experiment, a 9-min preincubation of PMN membranes with arachidonic acid and GTP-gamma-S followed by a 1-min contact with the cytosolic fraction led to a more marked activation than did preincubation of the cytosol with arachidonic acid and GTP-gamma-S for 9 min followed by a 1-min contact with membranes, suggesting the presence of a G-protein in the membrane fraction. In the absence of added cations, the reconstitution system exhibited a substantial oxidase activity which was totally prevented by ethylenediaminetetraacetic acid (EDTA). Mg2+ added at a concentration of 0.5-1 mM enhanced oxidase activation by about 30%, indicating that endogenous Mg2+ or other activating cations were sufficient to ensure 70% of maximal activation.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献