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1.
Temporal separation of nitrogen fixation and photosynthesis in the filamentous,non-heterocystous cyanobacterium Oscillatoria sp. 总被引:1,自引:0,他引:1
When growing in laternating light-dark cycles, nitrogenase activity (acetylene reduction) in the filamentous, non-heterocystous cyanobacterium Oscillatoria sp. strain 23 (Oldenburg) is predominantly present during the dark period. Dark respiration followed the same pattern as nitrogenase. Maximum activities of nitrogenase and respiration appeared at the same time and were 3.6 mol C2H4 and 1.4 mg O2 mg Chl a
-1·h-1, respectively. Cultures, adapted to light-dark cycles, but transferred to continuous light, retained their reciprocal rhythm of oxygenic photosynthesis and nitrogen fixation. Moreover, even in the light, oxygen uptake was observed at the same rate as in the dark. Oxygen uptake and nitrogenase activity coincided. However, nitrogenase activity in the light was 6 times as high (22 mol C2H4 mg Chl a
-1·h-1) as compared to the dark activity. Although some overlap was observed in which both oxygen evolution and nitrogenase activity occurred simultaneously, it was concluded that in Oscillatoria nitrogen fixation and photosynthesis are separated temporary. If present, light covered the energy demand of nitrogenase and respiration very probably fulfilled a protective function. 相似文献
2.
Mucor miehei lipase was immobilized on magnetic polysiloxane-polyvinyl alcohol particles by covalent binding. The resulting immobilized biocatalyst was recycled by seven assays, with a retained activity around 10% of its initial activity. Km and Vmax were respectively 228.3 M and 36.1 U mg of protein–1 for immobilized enzyme. Whereas the optimum temperature remained the same for both soluble and immobilized lipase (45 °C), there was a shift in pH profiles after immobilization. Optimum pH for the immobilized lipase was 8.0. Immobilized enzyme showed to be more resistant than soluble lipase when assays were performed out of the optimum temperature or pH. 相似文献
3.
Mesophyll cells isolated from Zinnia elegans L. cv. Canary Bird were cultured for 96 h in a liquid medium containing 0.1 mg l-1 -naphthaleneacetic acid and 1 mg l-1 benzyladenine in which both differentiation of tracheary elements (TE) and cell division were induced, or in a medium containing 0.1 mg l-1 -naphthaleneacetic acid and 0.001 mg l-1 benzyladenine, in which cell division was induced but TE differentiation was not. Lignification was found to occur only in the former medium, fairly synchronously after 76 h of culture, 5 h later than the onset of visible secondary wall thickening. Changes in the soluble phenolics were not correlated with TE differentiation. Of three important enzymes which have been reported to play a role in TE differentiation, the activity of phenylalanine ammonia-lyase (EC 4.3.1.5) in the TE-inductive culture was higher than that in the control culture between 72 and 96 h of culture, when TE differentiation progressed and lignin was synthesized actively. O-Methyltransferase (EC 2.1.1.6) activity was higher in the control culture than in the TE-inductive culture, indicating that this enzyme was not a marker enzyme of TE differentiation. The activities of peroxidases (EC 1.11.1.7), one extractable and the other nonextractable, with CaCl2 from the cell walls, reached peaks at 72 h (just before lignification) and 84 h of culture (active lignin synthesis), respectively, in the TE-inductive culture only, whereas the activity of soluble peroxidase showed a similar pattern of increase in the TE-inductive to the control culture. These results indicate that phenylalanine ammonia-lyase and peroxidase bound to the cell walls can be marker proteins for the differentiation of TE.Abbreviations OMT
O-methyltransferase
- PO
peroxidase
- PAL
phenylalanine ammonia-lyase
- TE
tracheary element(s) 相似文献
4.
Photosynthetic electron transport in thylakoids from cotton cultivars (Gossypium sp.) differing in tolerance to prometryn 总被引:1,自引:0,他引:1
A method to determine photosynthetic electron transport in thylakoid membranes is described for Gossypium barbadense (cv. Pima S-7) and G. hirsutum (cv. DP 5415). These cultivars differed markedly in tolerance to prometryn, a PS II inhibitor. The rates of photosynthetic electron transport obtained were 245 mole oxygen mg–1 chl h1. Plant age and leaf size influenced the activity of the thylakoid preparations. Thylakoids from leaves of plants 24 to 37 d and 50–70 mm in diameter had the highest activities; thylakoids from cotyledons, fully expanded leaves and young leaves had low activity. Thylakoids from both species had similar photosynthetic activities and I50's for prometryn, atrazine and diuron. Thus, tolerance to prometryn was not due to differential binding at D1 protein.Abbreviations PSII
photosystem II
- DAP
day after planting
- DQ
duroquinone
- DBMIB
dibromothymoquinone
- DMBQ
2,5-dimethyl-p-benzoquinone
- I50
concentration to inhibit reaction by 50%
- QA
quinone A
- QB
quinone B 相似文献
5.
Growth and nitrate uptake were studied on free-living and immobilizedChlorella vulgaris cells cultivated in medium containing different nitrate concentrations. First, the effect of nitrate concentrations on growth indicated that cells can live in the presence of high concentrations as high as 97 mM. Although no lethal effect on cells was observed such concentration a slow down in growth and a decrease in biomass produced was observed. The rate of nitrate uptake increased with the nitrate concentration in the medium. The maximum uptake rate was reached in first days of culture in both free-living and immobilized cells. The rate dropped more rapidly for cells growing in 2 mM nitrate than for cells growing in higher nitrate concentration. The maximum rate was very much the same for free-living and immobilized and was within the order of 0.45 to 0.57 g NO3 h–1 10–6 cells. Immobilization modified the changes of nitrate uptake rate for concentration higher than 2 mM. 相似文献
6.
Acetate uptake by strains of Synechococcus and Aphanocapsa in short experiments required light, and was strongly inhibited by m-dichlorocarbonyl cyanide phenylhydrazone and dichlorophenyl dimethyl urea. Acetate carbon was distributed in amino acids and in the acyl portion of lipids in the same way as during growth experiments when CO2 was available, but the reduced incorporation in the absence of CO2 was primarily into the lipid fraction. An apparent K
m for uptake by Synechococcus and for Aphanocapsa 6308 of 20 and 180 M at pH 7.4 was obtained; corresponding V
max values were 6 and 11 nmol x min-1 x mg protein-1. Uptake with Synechococcus was affected by pH, with affinity decreased and maximal rate increase with rising pH. Acetate uptake was not affected by propionate or butyrate when both were added at the same time, but a light and concentration dependent inhibition developed if suspensions were preincubated with propionate. Acetate carbon moved rapidly into acid insoluble material, but after 10–15 s 75% or more of the recovered intracellular counts were in acetyl CoA. Counts in this compound were reduced by preincubation with propionate.Kinetic measurements of acetyl CoA synthetase in fractionated cell extracts gave values for K
m of about 50 M for acetate, 5 mM for propionate, 100 M for CoA and 0.38 mM for ATP. The internal pool of free CoA was measured to be about 20 M, and was reduced by preincubation with propionate. This suggests that the activity of CoA-mediated reactions may be regulated by the availability of this cofactor.Abbreviations Used CCCP
m-Dichlorocarbonyl cyanide phenyl hydrazone
- DCMU
dichlorophenyl dimethyl urea
- TCA
trichloroacetic acid
- Tris
trishydroxymethyl amino methane
- HEPES
N-2-hydroxyethylpiperazine-N-2-ethane-sulfonic acid 相似文献
7.
A. R. Klein J. Breitung D. Linder K. O. Stetter R. K. Thauer 《Archives of microbiology》1993,159(3):213-219
Archaeoglobus fulgidus and Methanopyrus kandleri are both extremely thermophilic Archaea with a growth temperature optimum at 83°C and 98°C, respectively. Both Archaea contain an active N
5,N
10-methenyltetrahydromethanopterin cyclohydrolase. The enzyme from M. kandleri has recently been characterized. We describe here the purification and properties of the enzyme from A. fulgidus.The cyclohydrolase from A. fulgidus was purified 180-fold to apparent homogeneity and its properties were compared with those recently published for the cyclohydrolase from M. kandleri. The two cytoplasmic enzymes were found to have very similar molecular and catalytic properties. They differed, however, significantly with respect of the effect of K2HPO4 and of other salts on the activity and the stability. The cyclohydrolase from A. fulgidus required relatively high concentrations of K2HPO4 (1 M) for optimal thermostability at 90°C but did not require salts for activity. Vice versa, the enzyme from M. kandleri was dependent on high K2HPO4 concentrations (1.5 M) for optimal activity but not for thermostability. Thus the activity and structural stability of the two thermophilic enzymes depend in a completely different way on the concentration of inorganic salts. The molecular basis for these differences are discussed.Abbreviations H4MPT
tetrahydromethanopterin
- MFR
methanofuran
- CH3–H4MPT
N
5-methyl-H4MPT
- CH2=H4MPT
N
5,N
10-methylene-H4MPT
- CH2H4MPT
N
5,N
10-methenyl-H4MPT
- CHO–H4MPT
N
5 formyl-H4MPT
- CHO-MFR
formyl-MFR
- cyclohydrolase
N
5,N
10-methenyltetrahydromethanopterin cyclohydrolase
- MOPS
3-(N-morpholino) propane sulfonic acid
- TRICINE
N-tris (hydroxymethyl) methyl glycine
- 1 U=1
mol/min 相似文献
8.
Free-living Rhizobium trifolii MNF 1001 and cowpea Rhizobium MNF 2030 grown in chemostat culture under nitrogen limitation had high activities of an ammonium permease. In phosphate-limited, nitrogen-excess conditions, strains MNF 1001 and MNF 2030 retained 20% and 50%, respectively, of the ammonium uptake activity found in nitrogen-limited cells. Uptake in both strains was sensitive to azide, cyanide, carbonyl cyanide m-chlorophenyl hydrazone and 2,4-dinitrophenol. A gradient of ammonium concentration greater than 150-fold developed across the membrane within 20 min in cells of strain MNF 1001 grown under ammonia limitation. The pH optimum for ammonium uptake by N-limited cells of both MNF 1001 and MNF 2030 was around pH 7. The apparent K
m values for the ammonium permease in strains MNF 2030 and MNF 1001 were 3.9±1.6 M and 2.0±1.6 M respectively, and the V
max was 47±2.6 nmol min-1 (mg protein)-1 for MNF 2030 and 101±5.1 nmol min-1 (mg protein)-1 for MNF 1001. Isolated snake bean bacteroids of strain MNF 2030 capable of transporting succinate and l-glutamate had no detectable ammonium uptake activity. It therefore appears that the ammonium permeases in cells of these two strains are not as tightly regulated as in R. leguminosarum MNF 3841.Abbreviations CCCP
Carbonyl cyanide m-chlorophenyl hydrzone
- HEPES
N-Hydroxyethylpiperazine-N-2-ethanesulphonic acid 相似文献
9.
Several mutants and other variants of Myxococcus xanthus HP100 were obtained with differences in their sensitivity to carbenicillin and other penicillin derivatives. The specific activities of -lactamase in different resistant organisms varied from strain to strain but were consistently higher than in HP100. The relative molecular mass (M
r
) of the enzyme in M. xanthus HP100 was found to be 22,300. In certain carbenicillin resistant strains a second fraction of -lactamase activity of molecular weight 186,000 presumed to be an octamer of the other form was present. The enzyme was found in cell free extracts and also in culture supernatants of all carbenicillin resistant mutants but not in culture supernatants of strain HP100. In all the carbenicillin resistant mutants a part of the intracellular enzyme activity was released by osmotic shock and this activity may be periplasmic. The forms of the enzyme present in the culture supernatants and released by osmotic shock were monomeric. Carbenicillin resistance was not transferable between strains by conjugation. One resistance allele inhibited the transfer of the R factor Sa between myxococci.Non-standard abbreviations CS
CR sensitivity and resistance to carbenicillin
- C
u
R
C
S
R
unstable and stable resistance to carbenicillin 相似文献
10.
Cotton (Gossypium hirsutum) seeds and Sorghum vulgare caryopses are able to incorporate CO2 through a PEP-carboxylating enzyme (EC 4.1.1.38). The enzyme activity is optimal at pH 8.2 and is unaffected by ATP, GDP or acetyl CoA. The partially purified cotton enzyme is stimulated by inorganic phosphate with an apparent Km of 0.3 mM. The enzymes from both cultivars are inhibited by pyrophosphate, malate, and aspartate but not by succinate. Kinetic studies for Sorghum and cotton seed enzymes show apparent Km values for carbonate of 5 mM and 1.2 mM and for PEP of 36 M and 5 mM, respectively. The Vmax values are 90 and 3.3 nmol min-1 mg protein-1, respectively.A two-fold increase in the enzyme activity from cotton seeds occurs after 2 h under laboratory germination conditions after which the activity drops sharply to 1/3 of the original activity after 5 h imbibition. No such change was observed in Sorghum caryopses enzyme. A correlation between PEP-carboxylase activity and seed vigor in both cultivars was demonstrated.Abbreviations GOT
glutamicoxaloacetic-transaminase
- MDH
malic dehydrogenase-NADH2
- RH
relative humidity 相似文献
11.
José Edmundo Nava Saucedo Jean-Noël Barbotin Daniel Thomas 《Applied microbiology and biotechnology》1989,30(3):226-233
Summary The continuous production of gibberellic acid with immobilized mycelia of Gibberella fujikuroi was maintained over a hundred days in a tubular fixed-bed reactor. Free mycelium at the beginning of the storage phase was harvested from G. fujikuroi shake-flask culture and was immobilized by ionotropic gelation in calcium alginate beads.The continuous recycle production system consisted of a fixed-bed reactor, a container in which the culture medium was heated, stirred and aerated, and valves for sample withdrawal or reactant addition during the first 1320 h (55 days). A two-phase continuous extractor was then added for the last 960 hours (40 days). Free and immobilized mycelium shake-flask cultures with the same strain used in the continuous culture system were also realized to compare growth, maintenance and production parameters. The results show about the same gibberellic acid productivity in both free and immobilized mycelium shakeflask cultures: 0.384 and 0.408 mgGA3·gBiomass-1 ·day-1, respectively, whereas in the continuous system the gibberellic acid production is about twice as large for a similar biomass: 0.768 mgGA3·gBiomass-1·day-1. Several factors affecting the overall productivity of the immobilized systems were found to be: the quality and the quantity of mycelia in the biocatalyst beads and the immobilization conditions. 相似文献
12.
Dietrich Werner Arno Krotzky Regine Berggold Heidemarie Thierfelder Marianne Preiß 《Archives of microbiology》1982,132(1):51-56
Specific nitrogenase activity inAzospirillum brasilense ATCC 29145 in surface cultures under air is enhanced from about 50 nmol C2H4·mg protein-1·h-1 to 400 nmol C2H4 by the addition of 1 mM phenol. 0.5 and 2 mM phenol added increase the rate 5-fold and 4-fold. This enhancement effect is observed only between 2 and 3 days after inoculation, with only a small reduction of the growth of the cells by the phenol added. In surface cultures under 1% O2, nitrogenase activity is slightly reduced by the addition of 1–0.01 mM phenol. Utilization of succinate is enhanced during the period of maximum enhancement of nitrogenase activity by 60% by addition of 1 mM phenol. The cells did not produce14CO2 from [U-14C] phenol, neither in surface cultures nor in liquid cultures and less than 0.1% of the phenol was incorporated into the cells. A smaller but significant enhancement of nitrogenase activity by about 100% in surface cultures under air was found withKlebsiella pneumoniae K 11 after addition of 1 mM phenol. However, inRhizobium japonicum 61-A-101 all phenol concentrations above 0.01 mM reduced nitrogenase activity. With 1 mM phenol added activity was reduced to less than 10% with no effect on the growth in the same cultivation system. With thisRhizobium japonicum strain significant quantities of phenol (25 mol in 24 h by 2·1012 cells) were metabolized to14CO2, with phenol as sole carbon source. WithAzospirillum brasilense in liquid culture under 1% and 2% O2 in the gas phase, no enhancement of nitrogenase activity by phenol was noticed. 相似文献
13.
Human arylamine N-acetyltransferase 1 (NAT1) has been overexpressed in E. coli as a mutant dihydrofolic acid reductase (DHFR) fusion protein with a thrombin sensitive linker. An initial DEAE anion-exchange chromatography resulted in partial purification of the fusion protein. The fusion protein was cleaved with thrombin, and human rNAT1 was purified with a second DEAE column. A total of 8 mg of human rNAT1 from 2 l of cell culture was purified to homogeneity with this methodology. Arylamine substrate specificities were determined for human rNAT1 and hamster rNAT2. With both NATs, the second order rate constants (kcat/Kmb) for p-aminobenzoic acid (PABA) and 2-aminofluorene (2-AF) were several thousand-fold higher than those for procainamide (PA), consistent with the expected substrate specificities of the enzymes. However, p-aminosalicylic acid (PAS), previously reported to be a human NAT1 and hamster NAT2 selective substrate, exhibits 20-fold higher specificity for hamster rNAT2 (k cat/Kmb3410 M-1 s-1 ) than for human rNAT1 (kcat/Kmb 169.4 M-1 s-1 ). p-aminobenzoylglutamic acid (pABglu) was acetylated 10-fold more efficiently by human rNAT1 than by hamster rNAT2. Inhibition studies of human rNAT1 and hamster rNAT2 revealed that folic acid and methotrexate (MTX) are competitive inhibitors of both the unacetylated and acetylated forms of the enzymes, with KI values in 50–300 range. Dihydrofolic acid (DHF) was a much poorer inhibitor of human rNAT1 than of hamster rNAT2. The combined results demonstrate that human rNAT1 and hamster rNAT2 have similar but distinct kinetic properties with certain substrates, and suggest that folic acid, at least in the non-polyglutamate form, may not have an effect on human NAT1 activity in vivo. 相似文献
14.
Baterial lipase from Staphylococcus carnosus (pLipMut2) has been immobilized on various supports in order to determine a suitable immobilization technique in terms of activity and stability, when utilized for the hydrolysis of tributyrin. The hydrophobic materials PBA Eupergit and PBA Eupergit 250L prooved to be appropriate supports, when the enzyme was crosslinked with glutaraldehyde after adsorption. No desorption of the immobilized enzyme occured during operation. The pore size of the support has a strong effect on the activity but does not influence stability.The initial activity for immobilized and soluble lipase is found to follow the Arrhenius equation at low temperature, where mass transfer does not affect reaction kinetics. Activation energies for soluble and immobilized lipase were evaluated to be 21.7 kJ mol–1 and 60.8 kJ mol–1, respectively.Operational stability was studied in a packed bed recirculation reactor. Thermal desactivation followed first order kinetics with a half-life of 1340 h at 10°C. Model calculations for productivity showed, that optimal temperatures for high productivity are well below the temperature of maximal activity.List of Symbols
E
a
[kJ mol–1]
activation energy
-
E
d
[kJ mol–1]
activation energy of desactivation
-
H [–]
half-number
-
k
d
[h–1]
desactivation constant
-
k
d, [h–1]
constant
-
k
N
[–]
desactivation constant (number)
-
N [–]
number of runs
-
p [mol dm–3]
productivity
-
t [h]
time
-
t
0.5 [h]
half-life
-
T [K]
absolute temperature
-
V [U ml–1]
activity
- V(N) [Uml–1]
activity exhibited in the n-th run
-
V
s,O [U ml–1]
initial activity of supernatant
-
V
s, [U ml–1]
activity of supernatant after immobilization
-
V
O [U ml–1]
initial activity
-
V
[U ml–1]
constant
-
imm
[–]
activity yield
-
[ml ml–1]
ratio of volume of support to volume of supernatant
Financial support of this work by the Deutsche Forschungsgemeinschaft (SFB 145, A15) is gratefully acknowledged. 相似文献
15.
Ammonia reversibly inhibits the nitrogenase activity not only in purple nonsulfur bacteria but in purple (Thiocapsa roseopersicina) and green (Chlorobium limicola forma thiosulfatophilum) sulfur bacteria as well.The complete inhibition of nitrogenase activity (acetylene reduction) is observed about 30 s after addition of NH
4
+
(2.5×10-6 M) to cell suspensions. The pattern of ammonia inhibition of acetylene reduction in T. roseopersicina does not differ from the action of tetrabutylammonium and tetraphenylphosphonium (3 · 10-6-5·10-5 M) on nitrogenase activity of this bacterium.Simultaneously with the switch-off effect of NH
4
+
a considerable increase of ATP in cells of Rhodobacter sphaeroides and C. limicola f. thiosulphatophilum was observed. 相似文献
16.
The l-alanine dehydrogenase (ADH) of Anabaena cylindrica has been purified 700-fold. It has a molecular weight of approximately 270000, has 6 sub-units, each of molecular weight approximately 43000, and shows activity both in the aminating and deaminating directions. The enzyme is NADH/NAD+ specific and oxaloacetate can partially substitute for pyruvate. The K
m
app
for NAD+ is 14 M and 60 M at low and high NAD+ concentrations, respectively. The K
m
app
for l-alanine is 0.4 mM, that for pyruvate is 0.11 mM, and that for oxaloacetate is 3.0 mM. The K
m
app
for NH
4
+
varies from 8–133 mM depending on the pH, being lowest at high pH levels (pH 8.7 or above). Alanine, serine and glycine inhibit ADH activity in the aminating direction. The enzyme is active both in heterocysts and vegetative cells and activity is higher in nitrogen-starved cultures than in N2-fixing cultures. The data suggest that although alanine is formed by the aminating activity of ADH, entry of newly fixed ammonia into organic combination does not occur primarily via ADH in N2-fixing cultures of A. cylindrica. Ammonia assimilation via ADH may be important in cultures with an excess of available nitrogen. The deaminating activity of the enzyme may be important under conditions of nitrogen-deficiency.Abbreviations ADH
alanine dehydrogenase
- DEAE
diethylamino ethyl cellulose
- EDTA
ethylenediamine tetraacetic acid
- GDH
glutamic dehydrogenase
- GS
glutamine synthetase
- GOT
aspartate-glutamate aminotransferase
- NAD+
nicotinamide adenine dinucleotide
- NADH
reduced nicotinamide adenine dinucleotide
- NADP+
nicotinamide adenine dinucleotide phosphate
- NADPH
reduced nicotinamide adenine dinucleotide phosphate
- SDS
sodium dodecyl sulphate
- Tris
tris(hydroxymethyl) aminomethane 相似文献
17.
The free-living cyanobacterium Anabaena variabilis showed a biphasic pattern of 14CH3NH
3
+
uptake. Initial accumulation (up to 60 s) was independent of CH3NH
3
+
metabolism, but long-term uptake was dependent on its metabolism via glutamine synthetase (GS). The CH3NH
3
+
was converted into methylglutamine which was not further metabolised. The addition of l-methionine-dl-sulphoximine (MSX), to inhibit GS, inhibited CH3NH
3
+
metabolism, but did not affect the CH3NH
3
+
transport system.NH
4
+
, when added after the addition of 14CH3NH
3
+
, caused the efflux of free CH3NH
3
+
; when added before 14CH3NH
3
+
, NH
4
+
inhibited its uptake indicating that both NH
4
+
and CH3NH
3
+
share a common transport system. Carbonylcyanide m-chlorophenylhydrazone and triphenyl-methylphosphonium both inhibited CH3NH
3
+
accumulation indicating that the transport system was -dependent. At pH 7 and at an external CH3NH
3
+
concentration of 30 mol dm-3, A. variabilis showed a 40-fold intracellular accumulation of CH3NH
3
+
(internal concentration 1.4 mmol dm-3). Packets of the symbiotic cyanobacterium Anabaena azollae, directly isolated from the water fern Azolla caroliniana, also showed a -dependent NH
4
+
transport system suggesting that the reduced inhibitory effect of NH
4
+
on nitrogenase cannot be attributed to the absence of an NH
4
+
transport system but is probably related to the reduced GS activity of the cyanobiont.Abbreviations CCCP
carbonylcyanide m-chlorophenylhydrazone
- GS
glutamine synthetase
- HEPES
4-(2-hydroxyethyl)-1-piperazine ethanesulphonic acid
- MSX
l-methionine-dl-sulphoximine
-
membrane potential
- pH
transmembrane pH difference
- TPMP+
triphenylmethylphosphonium 相似文献
18.
Ion stimulation and some other properties of an ATPase activity associated with vacuoles isolated from storage roots of red beet (Beta vulgaris L.) have been determined. The ATPase had a specific requirement for Mg2+ and in the presence of Mg2+ it was stimulated by salts of monovalent cations. The degree of stimulation by monovalent salts was influenced mainly by the anion and the order of effectiveness of the anions tested was Cl->HCO
3
-
>Br->malate>acetate>SO
4
2-
. For any given series of anions the magnitude of the stimulation obtained was influenced by the accompanying cation (NH
4
+
Na+>K+). This cation effect was abolished by 0.01% (v/v) Triton X-100 and it is suggested that it is the result of different permeabilities of membrane vesicles to the cations. There was no evidence of synergistic stimulation of the ATPase by mixtures of Na+ and K+. KCl- and NaCl-stimulation was maximal with salt concentrations in the range 60–150 mM. The true substrate of the enzyme was shown to be MgATP. It was shown that KCl stimulation was the result of an increase in Vmax rather than a change in the affinity of the enzyme for MgATP. The ATPase was inhibited by N,N-dicyclohexylcarbodiimide, diethylstilbestrol, mersalyl and KNO3 but other inhibitors tested (azide, oligomycin, orthovanadate, K3[Cr(oxalate)6] and ethyl-3-[3-dimethylaminopropyl]carbodiimide) were without effect or caused only partial inhibition at the highest concentration tested. The ATPase activity was equally distributed between pellet and supernatant fractions obtained after the subfractionation of vacuoles but the properties of the ATPase in each fraction were the same. It is suggested that beet vacuoles possess only one ATPase. The properties of the ATPase are compared with those of ATPases associated with other plant membranes and organelles and its possible role in transport at the tonoplast is discussed.Abbreviations ATPF
free ATP
- ATPT
total ATP
- BSA
bovine serum albumen
- DCCD
N,N-dicyclohexylcarbodiimide
- DES
diethylstilbestrol
- DNP
2,4-dinitrophenol
- EDAC
ethyl-3-(3-dimethylaminopropyl)carbodiimide
- Km
apparent Michaelis constant
- MgATP
complex of Mg2+ and ATP
- Mg
F
2+
free Mg2+
- Mg
T
2
total Mg2+
- MES
2-(N-Morpholino)ethanesulphonic acid
- Na2EDTA
disodium ethylenediaminetetraacetic acid
- NEM
N-ethylmaleimide
- Pi
inorganic phosphate
- TCA
trichloroacetic acid
- Tris
tris(hydroxymethyl)methylamine
- Vmax
maximum velocity 相似文献
19.
Putrescine carbamoyltransferase (PutCT) has been postulated to function in the synthesis of putrescine (Put) from an N-carbamoylputrescine (NCPut) intermediate in plants. In pea, PutCT activity was associated entirely with ornithine carbamoyltransferase (OCT) protein, which was purified to homogeneity using an immobilized transition-state analog inhibitor (
N-(phosphonacetyl)-L-ornithine). No evidence for a separate PutCT enzyme, similar to that in Streptococcus [15], or PutCT activity associated with a putrescine synthase-type multifunctional enzyme [13] was found. OCT carried out the carbamoylation of Put and other diamine and polyamine substrates inefficiently and at non-physiological pH (Put carbamoylation: pH 10.8 optimum, Vmax 0.11 kat/mg protein, Km=6.7 mM for Put and 1.0 mM for carbamoyl-P), when compared with ornithine carbamoylation (pH 8.5 optimum, Vmax=313.9 kat/mg protein, Km=4.4 mM for ornithine and 0.6 mM for carbamoyl-P). Different subcellular compartmentation of PutCT activity (chloroplast) and the NCPut substrate (cytosol), coupled with a thermodynamically-unfavorable reverse reaction (i.e., Put synthesis from NCPut), suggest that the OCT-associated PutCT activity does not significantly contribute to in vivo Put synthesis in plants.Abbreviations ADC
Arginine Decarboxylase
- Agm
Agmatine
- AIH
Agmatine Iminohydrolase
- Cad
Cadaverine
- Cit
Citrulline
- CP
Carbamoyl-P
- Dap
1,3-Diaminopropane
- Dns-
Dansyl-
- NCPut
N-Carbamoylputrescine
- OCT
Ornithine Carbamoyltransferase
- Put
Putrescine
- PutCT
Putrescine Carbamoyltransferase
- Spd
Spermidine
- Spm
Spermine 相似文献
20.
G. M. Scarpa F. Pupilli F. Damiani S. Arcioni 《Plant Cell, Tissue and Organ Culture》1993,35(1):49-57
Seventeen ecotypes of the wild species Medicago polymorpha adapted to a Sardinian (Italy) environment have been evaluated for their response to tissue culture. The accession Samughero-Albi was the more respondent for callus induction and, together with Usassai, showed the highest regeneration capacity on media containing 1 mg l-1 2iP and 0.1 mg l-1 IAA. The morphogenetic response was also affected by the explant source. The hypocotyl-derived-calli were the best regenerating tissues. Regenerated plantlets were difficult to root and it was possible to obtain plants with a well developed root system only after 5–7 weeks of culture on media containing 2iP and IAA both at 0.2 mg l-1. Mesophyll cells were the best protoplast yielding source but only those isolated from roots were able to divide and to regenerate plants. Results are discussed in relation to the genotype specificity for the morphogenetic response and the feasibility of using M. polymorpha in the somatic hybridization with M. sativa.Abbreviations NAA
-naphthaleneacetic acid
- 6-BAP
6-benzylaminopurine
- 2,4-d
2,4-dichlorophenoxyacetic acid
- 2iP
N6-2-isopentenyl-adenine
- IAA
indole-3-acetic acid
- GA3
gibberellic acid
- GFMS
growth regulator free MS medium
- Prol
proline
- Malt
maltose 相似文献