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1.
胆碱脱氢酶的动力学性质 总被引:1,自引:0,他引:1
本文对增溶胆碱脱氢酶的稳态初速度及产物抑制动力学做了。底物胆碱和PMS的相互影响:变化一个底物的浓度,另一个底物的Km及Vmax均变化。该酶的产物三甲胺 地 制表现为对底物胆碱非竞争性而地PMS竞争性,在胆碱饱和的情况下,三甲胺乙醛对酶的抑制仍表现为对PMS竞争性。这些结果表明增溶胆碱脱氢酶的催化机制搂双底物双产物乒乓机制。1-PC与9-AC对增溶胆碱脱氢酶均有抑制作用,且均为混和型抑制,K1分别 相似文献
2.
酵母醇脱氢酶ADHI的纯化及动力学研究 总被引:3,自引:1,他引:2
本文报道了啤酒酵母醇脱氢酶组成型同工酶ADHI的快速高效的纯化方法。通过活性蓝色染料柱亲和层析的方法将该酶纯化至电泳均一,收率达47%,对该酶的产物抑制及端点抑制动力学研究结果支持Wratten,Cleland提出的序列有序机制。 相似文献
3.
Germinating seeds with the highest specific activity (24 hour germination) were used for isolation of alcohol dehydrogenase, ADH, from rape (Brassica napus L. cv. T?ebi?ská). The rape ADH was purified by fractionation with ammonium sulphate, desalting on Sephadex G 25, chromatography on DEAE cellulose and gel filtration on Sephadex G 150. Using this isolation procedure, enzyme with a specific activity 85.6 times higher than that of the crude extract was obtained. The molecular weight of the enzyme obtained is 66.000. The enzyme is a metallo-enzyme containing sulfhydryl groups as evidence by the inhibitory effect of chelating compounds and thiol reagents. The optimum pH for the oxidation of ethanol is 8.5 and for reduction of acetaldehyde 7.0. The enzyme exhibits a relatively wide substrate specificity towards alcohols. Dimethyl-sulphoxide (DMSO), some amides and oximes and some intermediates of the carbohydrate metabolism act as ADH inhibitors, ATP as analogue of NAD also exhibits an inhibitory effect. The inhibitory effect of heterocyclic substances (pyrazol, imidazol, pyridine) is similar to the effect on liver alcohol dehydrogenase. 相似文献
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5.
The process of antigen presentation is not well understood. We screened for drugs that distinguish presentation of allogeneic class 2 antigens and exogenous antigens. When spleen cells were used as antigen presenting cells (APC), leupeptin and antipain preferentially inhibited allogeneic class 2 presentation, while they did not affect presentation of exogenous antigen and T cell growth. In contrast, they inhibited both presentations when spleen adherent cells (SAC) were used as APC. Our results suggest that SAC (mainly macrophages) and splenic B cells use different pathways to present exogenous antigens and that pathways to present allogeneic class 2 molecules are similar. 相似文献
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7.
Changing Kinetic Properties of Fructose-1,6-bisphosphatase from Pea Chloroplasts during Photosynthetic Induction
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After dark-light transitions, there is a delay in photosynthetic CO2 fixation by isolated pea chloroplasts in the range of some minutes. In order to assess the physiological significance of light modulation of enzyme activity in the control of induction, we made estimates of the kinetic parameters of fructose-1,6-bisphosphatase immediately upon release from pea chloroplasts in the dark and after illumination for various time periods. The Michaelis constant for fructose-1,6-bisphosphate decreased and maximal velocities increased during induction. It seems likely that light activation of this enzyme is one of the factors contributing to the overcoming of the lag period in photosynthetic CO2 fixation. 相似文献
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Properties of a Large Complex with NADH Dehydrogenase Activity from Barley Thylakoids 总被引:4,自引:0,他引:4
Cuello Juan; Quiles Maria Jose; Albacete Maria Eugenia; Sabater Bartolome 《Plant & cell physiology》1995,36(2):265-271
When assays for NAD(P)H-ferricyanide oxidoreductases were performed,activities specific for NADH (0.23 unit (mg protein)1)and NADPH (0.68 unit (mg protein)1) were detected inchloroplasts isolated from leaves of barley (Hordeum vulgareL.). Activities of chloroplast NADH- and NADPH-ferricyanideoxidoreductase were 5-fold and 25-fold higher, respectively,than the maximum activity that could be attributed to mitochondrialcontamination. Moreover, most of the chloroplast NADH-ferricyanideoxidoreductase (60 to 80%) was solubilized by deoxycholate (DOC)from thylakoids as a single, high-molecular-mass complex thatwas distinguishable from the mitochondrial complex by its lowerelectrophoretic mobility in 3% polyacrylamide, as revealed byreduction of nitro blue tetrazolium (NBT) in the presence ofNADH or NADPH on gels after electrophoresis. The stroma yieldeda single band of a dehydrogenase (66 kDa) that used NADH asits electron donor. Several NADPH-dependent activities weredetected after electrophoresis of the stromal fraction. Moreover,chloroplast-specific activities could be distinguished frommitochondrial activities on the basis of the specificity ofthe donor and the acceptor of electrons, the dependence of theactivities on pH, and the sensitivity to various inhibitors.Km values for NADH (26 µM) and NADPH (75 µM) werein the same range as those of mitochondrial activities. Mostof the NADPH-dependent activity probably corresponds to thechloroplast ferredoxin-NADP+ oxidoreductase. The possibilityis discussed that thylakoid NADH dehydrogenase(s) might be theproduct of chloroplast ndh genes and that this activity is involvedin chlororespiration. (Received April 25, 1994; Accepted December 5, 1994) 相似文献
10.
6-Phosphogluconate dehydrogenase has been purified from human brain to a specific activity of 22.8 U/mg protein. The molecular weight was 90,000. At low ionic strengths enzyme activity increased, due to an increase in Vmax and a decrease in Km for 6-phosphogluconate, and activity subsequently decreased as the ionic strength was increased (above 0.12). Both 6-phosphogluconate and NADP+ provided good protection against thermal inactivation, with 6-phosphogluconate also providing considerable protection against loss of activity caused by p-chloromercuribenzoate and iodoacetamide. Initial velocity studies indicated the enzyme mechanism was sequential. NADPH was a competitive inhibitor with respect to NADP+, and the Ki values for this inhibition were dependent on the concentration of 6-phosphogluconate. Product inhibition by NADPH was noncompetitive when 6-phosphogluconate was the variable substrate, whereas inhibition by the products CO2 and ribulose 5-phosphogluconate and NADP+ were varied. In totality these data suggest that binding of substrates to the enzyme is random. CO2 and ribulose 5-phosphate are released from the enzyme in random order with NADPH as the last product released. 相似文献
11.
Alcohol Dehydrogenase of Apple 总被引:3,自引:0,他引:3
The alcohol dehydrogenase prepared from apple (Malus domesticaBorkh.) possesses both NADH and NADPH-linked activities, whenassayed with acetaldehyde as substrate. The pyridine nucleotidesbind to the same catalytic site on the enzyme. The alcohol dehydrogenasecan also catalyse the reduction of C3C6 aldehydes witheither NADH or NADPH as cofactor. 相似文献
12.
6-Phosphogluconate dehydrogenase (6PG) was purified from rat small intestine with 36% yield and a specific activity of 15 U/mg.
On SDS/PAGE, one band with a mass of 52 kDa was found. On native PAGE three protein and two activity bands were observed.
The pH optimum was 7.35. Using Arrhenius plots, Ea, ΔH, Q10 and Tm for 6PGD were found to be 7.52 kcal/mol, 6.90 kcal/mol, 1.49 and 49.4°C, respectively. The enzyme obeyed “Rapid Equilibrium
Random Bi Bi” kinetic model with Km values of 595 ± 213 μM for 6PG and 53.03±1.99 μM for NADP. 1/Vm versus 1/6PG and 1/NADP plots gave a Vm value of 8.91±1.92 U/mg protein. NADPH is the competitive inhibitor with a Ki of 31.91±1.31 μM. The relatively small Ki for the 6PGD:NADPH complex indicates the importance of NADPH in the regulation of the pentose phosphate pathway through G6PD
and 6PGD. 相似文献
13.
The final activity of the alcohol dehydrogenase (E.C.1.1.1.1, abbreviated ADH) from germinating pea, isolated by fractionating with ammonium sulphate, chromatography on DEAE cellulose and gel filtration, was 80,000, from bean 25,000 and from lentil 13,500 units per mg protein. Molecular weights of the ADHs are close to each other: pea and bean ADH 60,000, lentil ADH 70,000. The Km values are mutually similar with three enzymes, i.e. of the order of 10−4M for NAD and 10−2M for ethanol. The pH optima lie in the alkaline region. These enzymes catalyse oxidation of a number of monovalent alcohols. At temperatures above 60°C the enzymes are thermally unstable. Stability is enhanced slowly by ethanol but not by NAD. Pyrazol, imidazol and pyridine inhibit plant ADH similarly to the enzyme from horse liver. There is a similarity between plant alcohol dehydrogenases and animal and yeast enzymes. 相似文献
14.
Semenihina A. Popova T. Schnarrenberger C. Pinheiro De Carvalho M.A.A. 《Photosynthetica》2001,39(2):299-304
The cytosolic and chloroplastic isoforms of glucose-6-phosphate dehydrogenase (G6PDH) were separated and purified from barley leaves (Hordeum vulgare L.). In etiolated leaves, only the cytosolic isoform was expressed. The molecular mass of the cytosolic enzyme, G6PDH1, was 112±8 kDa and that of the chloroplast enzyme, G6PDH2, was 136±7 kDa. The Km values for glucose-6-phosphate and NADP were 0.133 and 0.041 mM for G6PDH1, and 0.275 and 0.062 mM for G6PDH2, respectively. The pH optimum was 8.2 for G6PDH1 and 7.8 for G6PDH2. The enzyme is absolutely specific for NADP. NADPH is a competitive inhibitor of the G6PDH1 in respect to glucose-6-phosphate (G6P) and NADP (Ki = 0.050 and 0.025 mM, respectively). NADPH is a competitive inhibitor of the G6PDH2 in respect to NADP (Ki = 0.010 mM), but a non-competitive inhibitor in respect to the G6P. ADP, AMP, UTP, NAD, and NADH had no effect on the activity of G6PDH. ATP inhibited the G6PDH2 activity. 相似文献
15.
The study of ß-glycerophosphatase activity in cell-wallpreparations and in excised root tips from barley seedlingssupports the view that the former, which constitutes about 20per cent of the activity of the whole homogenate, representsthe fraction located at the surface of the roots in vivo. Theactivities of the cell-wall suspension and intact roots arevirtually identical, and further show identical relations topH, substrate concentration (Km), and competitive inhibitionby molybdate and inorganic phosphate (Ki). The enzyme must thereforebe freely exposed to the external solution without any permeabilitybarrier separating it from either substrate or inhibitors. Theabsence of any lag phase in the hydrolysis in excised root tipssuggests that the surface enzyme may be limited to the outermostlayers of the root. The solubilization of some of the activityof the cell-wall preparation by treatment with sodium chlorideand ammonium sulphate suggest that surface activity may havebeen lost from these preparations rather than adsorbed duringhomogenization and extraction. The Km and pH-activity curveof the supernatant activity remaining after centrifugation ofthe cell-wall fraction indicate that only this enzyme and noother detectable glycerophosphatase exists in the roots. 相似文献
16.
The Alcohol Dehydrogenase Polymorphism in Drosophila Melanogaster: Fitness Measurements and Predictions under Conditions with No Alcohol Stress 总被引:1,自引:1,他引:0
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The relative fitnesses of the different Adh genotypes under normal laboratory conditions and in the absence of alcohol stress were estimated in Drosophila melanogaster according to Prout's method. The larval component (viability) did not reveal fitness differences between the genotypes but for the adult component significant differences were observed. The female adult component (fecundity) showed an overdominant pattern: both homozygous genotypes showed a relative fitness significantly lower than the heterozygous genotype. For the male adult component (virility) also differences were observed. The homozygous SS genotype showed a lower relative fitness than the other two genotypes. Predictions for gene frequency changes based on the estimated fitness values do show a reasonably good correspondence with frequency changes actually observed in a number of experimental cage populations and indicate a globally stable equilibrium around a frequency of the F allele of 0.40-0.70. The relevance of these fitness estimates, obtained under conditions with no alcohol stress, for the explanation of the Adh polymorphisms observed in nature is discussed. 相似文献
17.
Arakawa Keita; Mizuno Katsuhiko; Kishitani Sachie; Takabe Tetsuko 《Plant & cell physiology》1992,33(7):833-840
The changes in the level of the protein for betaine aldehydedehydrogenase, which catalyzes the last step in the synthesisof glycinebetaine, were analyzed with antiserum raised againstSDS-denatured betaine aldehyde dehydrogenase from spinach. Inbarley leaves, the levels of betaine aldehyde dehydrogenaseprotein were found to be enhanced by the addition of 200 mMNaCl to the growth medium. These changes in the level of theenzyme protein corresponded to those in the activity of theenzyme, as described in our previous study (Arakawa et al. 1990).The extent of this enhancement was reduced when barley plantswere relieved from salt stress. An increase in the level ofthe protein was also induced by water stress, such as the withholdingof water or the addition of polyethylene glycol 6000. Betainealdehyde dehydrogenase protein was detected in etiolated leavesand roots, as well as in green leaves. In etiolated leaves,the level of betaine aldehyde dehydrogenase protein was notaffected by salt stress.
1 This work was supported by a grant from the Bio-Media Projectof the Japanese Ministry of Agriculture, Forestry and Fisheries(BMP92-III-l-1). 相似文献
18.
从食醋生产企业的醋醅中采集样品,以乙醇为唯一碳源,用碳酸钙透明圈平板法分离出185株菌株,然后以产酸量和耐乙醇能力为标准,瓶发酵选育出20株ADH产酶菌株;A5-2产酸量为49.85 g/L,耐乙醇能力强,A5-2的菌种形态学和16S rDNA序列分析初步鉴定为巴斯德醋酸杆菌( Acetobacter pasteurianus);A5-2乙醇脱氢酶酶学性质研究表明:最适作用温度和pH分别为45℃和pH 4.0,具有一定的耐热性和良好的耐酸碱性;A5-2乙醇脱氢酶粗酶制备条件为硫酸铵饱和度70%~80%,回收率84%。 相似文献
19.
Losses of alcohol, which had accumulated under anaerobic conditions,occurred during the germination of several species of seedswhich could not be attributed to the volatility of the alcohol.It is suggested that utilization of the alcohol by the seedsmay occur. From the seeds, an active alcohol dehydrogenase,which is mainly confined to the cotyledons in pea seeds, canbe extracted. The activity of the enzyme decreases as the cotyledonsgrow older during germination. The properties of the enzymehave been investigated. 相似文献
20.
Changing Activity of Glucose-6-Phosphate Dehydrogenase from Pea Chloroplasts during Photosynthetic Induction 总被引:2,自引:2,他引:0
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Light inactivation of glucose 6-phosphate dehydrogenase is rapid and occurs before photosynthetic O2 evolution is measureable in intact chloroplasts. Likewise, dark activation is rapid. The major light induced change in the kinetic parameters of glucose 6-phosphate dehydrogenase is in maximal velocity. 相似文献