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1.
The utilization of sugars and starch as carbon sources by sugarcane cell suspension cultures 总被引:1,自引:0,他引:1
Best growth of sugarcane cell suspensions on sugar substrateswas obtained with raffinose, followed closely by sucrose. Growthon glucose, fructose, cellobiose, melibiose and trehalose wasslightly below that on sucrose. Pentoses have been reportedpreviously not to be utilized by plant tissues in culture-ribosesupported growth of the sugarcane cells. Starch supported growthas well as did sucrose. This support is due to the secretionof an -amylase by the sugarcane cells in culture.
1Published with the approval of the Director as Paper No. 239in the Journal Series of the Experiment Station, Hawaiian SugarPlanters' Association, Honolulu, Hawaii, U.S.A. (Received October 3, 1969; ) 相似文献
2.
In response to in situ dark modulation, or in vitro ATP preincubationof higher plant nitrate reductase, Mg2+ inhibits NADH:nitratereductase activity but not MV:nitrate reductase activity incrude extracts. Also for the purified enzyme the complete NADH:nitratereductase activity is inhibited by Mg2+, but not the partialMV:nitrate reductase or Cyt c reductase activities. (Received October 13, 1993; Accepted January 24, 1994) 相似文献
3.
Funkhouser Edward A.; Gewitz Hans-Siegfied; Vennesland Birgit 《Plant & cell physiology》1983,24(8):1565-1568
During induction of nitrate reductase in Chlorella vulgaris,synthesis of the precursor, demolybdo cytochrome c reductase,exceeds the synthesis of active enzyme. Evidence is also presentedwhich shows that the purification procedure of Funkhouser etal. [(1980) Plant Physiol. 65: 939] separates demolybdo cytochromec reductase from active nitrate reductase.
1Supported in part by a grant to B. V. from the Deutsche Forschungsgemeinschaftand a contribution of the Texas Agricultural Experiment Station. (Received July 27, 1983; Accepted September 13, 1983) 相似文献
4.
Mahan James R.; Sherman Timothy D.; Funkhouser Edward A. 《Plant & cell physiology》1988,29(4):735-737
The thermal dependence of two of the reactions catalyzed bythe nitrate reductase from Chlorella vulgaris was determined.The activation energies for NADH:nitrate oxidoreductase (EC1.6.6.1
[EC]
) and NADH:Cytochrome c oxidoreductase (EC 1.6.99.3
[EC]
)are 42.1 kJ?mol1 and 21.5 kJ?mol1, respectively.Since the thermal dependency of the two enzymes is different,ratios of the activities will vary with temperature. The importanceof both rigorous thermal control during nitrate reductase assaysas well as the need to specify the temperature at which theratio of activities for the enzyme are clearly established.
1Present Address: Cropping Systems Research Laboratory, USDA-ARS,Route 3, Box 215, Lubbock, TX 79401, U.S.A. (Received November 25, 1987; Accepted March 2, 1988) 相似文献
5.
Expression in Escherichia coli of Cytochrome c Reductase Activity from a Maize NADH:Nitrate Reductase Complementary DNA 总被引:6,自引:2,他引:4
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Campbell WH 《Plant physiology》1992,99(2):693-699
A cDNA clone was isolated from a maize (Zea mays L. cv W64A×W183E) scutellum λgt11 library using maize leaf NADH:nitrate reductase Zmnr1 cDNA clone as a hybridization probe; it was designated Zmnr1S. Zmnr1S was shown to be an NADH:nitrate reductase clone by nucleotide sequencing and comparison of its deduced amino acid sequence to Zmnr1. Zmnr1S, which is 1.8 kilobases in length and contains the code for both the cytochrome b and flavin adenine dinucleotide domains of nitrate reductase, was cloned into the EcoRI site of the Escherichia coli expression vector pET5b and expressed. The cell lysate contained NADH:cytochrome c reductase activity, which is a characteristic partial activity of NADH:nitrate reductase dependent on the cytochrome b and flavin adenine dinucleotide domains. Recombinant cytochrome c reductase was purified by immunoaffinity chromatography on monoclonal antibody Zm2(69) Sepharose. The purified cytochrome c reductase, which had a major size of 43 kilodaltons, was inhibited by polyclonal antibodies for maize leaf NADH:nitrate reductase and bound these antibodies when blotted to nitrocellulose. Ultraviolet and visible spectra of oxidized and NADH-reduced recombinant cytochrome c reductase were nearly identical with those of maize leaf NADH:nitrate reductase. These two enzyme forms also had very similar kinetic properties with respect to NADH-dependent cytochrome c and ferricyanide reduction. 相似文献
6.
The enzyme nitrate reductase could not be detected in leaf tissuesof cauliflower plants grown in sterile cultures with glutamicacid or ammonium sulphate if nitrate was absent. Excised leaftissues from these plants formed the enzyme for several hoursat a steady rate when infiltrated with nitrate. Plants starvedof nitrate for short periods lost enzyme activity which wasrestored in excised tissues upon infiltration with nitrate butnot with ammonium sulphate or nitrite. Molybdenum-deficientplants grown with nitrate also lacked enzyme activity whichwas restored in excised tissues after infiltration with molybdenum.Both nitrate and molybdenum were required to produce maximalrates of enzyme formation in excised tissues of plants grownwith ammonium sulphate and no molybdenum. Apparent Michaelisconstants for nitrate and molybdenum were found to be about10-5 and 10-7 respectively. The capacity of excised tissuesto respond to the inducer varied with their age and leaf positionon the plant and was exercised under conditions where growthwas unlikely. Increases in specific activities were similar.There was no evidence of a lag in response to nitrate or molybdenumwith tissues of plants grown with ammonium sulphate or glutamicacid in sterile cultures but lag periods were observed withtissues from plants deprived of nitrate. Cell-free preparationswere unable to respond to either factor. The results are interpretedas evidence for induced enzyme formation in vivo in responseto the substrate or the constituent metal. 相似文献
7.
The Occurrence of Nitrate Reduction in the Leaves of Woody Plants 总被引:13,自引:1,他引:12
Nitrate reductase activities greater than 02 µmol h1g1 f. wt, measured by an in vivo assay, occurred in 41per cent of a large sample (555 species) of woody plants. Ifseveral taxonomic groups (Gymnosperms, Ericaceae and Proteaceae)with consistently low activities were discounted activitiesgreater than 02 µmol h1 g1 f. wt occurredin 73 per cent of the species. This compares with 93 per centin herbaceous species, suggesting that leaf nitrate reductionis of common occurrence in woody plants. In a small sample ofspecies leaf nitrate reductase activity correlated with nitrateconcentration in the xylem sap. Low activities occurred consistentlyin the Gymnosperms, Ericaceae and Proteaceae. Feeding cut shootsof representatives of these groups with nitrate caused inductionof leaf nitrate reductase activity in the Gymnosperms and Proteaceae,but only limited induction in the Ericaceae. The Ericaceae,with the exception of two species, had low activities and lownitrate reductase inducibility. Root assimilation may predominatein the Gymnosperms and Proteaceae. It is suggested that nitratereduction generally occurs in the leaves of trees from a varietyof plant communities and that this may be related to the lowerenergy cost of leaf, as opposed to root, nitrate assimilation. Nitrate reductase, trees and shrubs, leaves, nitrate assimilation, nitrate translocation, nitrate reductase induction, energy cost, plant ecology 相似文献
8.
Spinach plants grown without molybdenum lack nitrate reductaseand when plants are deprived of nitrate existing activity islost. Transfer of molybdenum-deficient plants to a solutioncontaining (NH4)299MoO4) or nitrate-starved plants to NaNO3solution induced enzyme activity in 24 hr. After purificationby selective adsorption, precipitation and disc electrophoresis,the protein from molybdenum-deficient plants given 99Mo showedradioactivity only where nitrate reductase was revealed on theacrylamide gel. Molybdenum was similarly selectively concentratedinto the enzyme as a result of induction by nitrate in plantsgrown with sub-optimal molybdenum supply in order to minimizeeffects of isotope dilution on measurement of 99Mo incorporation. There was no exchange in vitro between 99Mo and purified activeenzyme in the resting state over 18 hr at 4°C, or with functioningenzyme held at room temperature for 24 hr. There was evidenceeither for possible in vivo exchange of 99Mo andenzyme boundMo or for slight synthesis of fresh enzyme under conditionsof net loss of enzyme in nitrate starved plants. Five NADH2 and two NADPH2 reactive diaphorases which could beseparated by electrophoresis were present in extracts. Onlyone of these having strong NADH2 and weak NADPH2 activity wasdirectly associated with nitrate reductase. The same complexalso showed the only benzyl viologen (BV.) reactive nitratereductase. Nitrate reductase in spinach is therefore considered to be amolybdenum-dependant and molybdenum-containing protein in whichNADH2 (with weak NADPH2) and BVelectron donor functions anddiaphorase/reductase activities remain closely associated duringpurification and electrophoresis. The techniques provide a simple means for the production andpurification of enzyme containing radioactively labelled Moapplicable to investigations on the structure of the enzyme. (Received January 16, 1971; ) 相似文献
9.
Hydrosulfite-reduced FMN served as an electron donor for nitratereductase purified from broad bean leaves. FMN was successfullyreplaced with BV. The flavine nucleotide nitrate reductase hadits pH optima at about 7.8 with phosphate buffer and at about7.4 with Tris-HCl buffer. The Km's for nitrate and FMN were3.7 ? 104 M and 3.7 ? 105 M, respectively. NADH2: nitrate reductase activity was completely inhibited by0.1 mM p-CMB, whereas FMNH2: nitrate reductase activity wasnot. Inhibited activity was restored by the addition of cysteine.A sulfhydryl enzyme is involved in the NADH2: nitrate reductasesystem but not in the FMNH2 : nitrate reductase system. NADH2and FMNH2 probably feed electrons into the electron transportchain at different sites. The nitrate reductase preparationhad an NADH2-specific diaphorase activity which was almost completelyinhibited by 0.1 mM p-CMB. The NADH2-specific diaphorase mayform the sulfhydryl enzyme which mediates electron transferbetween NADH2 and nitrate. (Received May 6, 1969; ) 相似文献
10.
Assimilatory nitrate reductase (NR) was solubilized by acetonetreatment from Plectonema boryanum and was purified 7,700-foldby heat treatment, ammonium sulfate fractionation and chromatographyon DEAE-Sephacel and Sephadex G-150. Purified NR had a specificactivity of 85 µmol NO2 formed min1 mg1protein. The enzyme retained both ferredoxin (Fd)- and methylviologen (MV)-linked NR activities throughout the purificationprocedure. Molecular weight was 80,000. The pH optimum was 10.5in the MV-assay and 8.5 when assayed with enzymatically reducedFd as the electron donor. Apparent Km values for nitrate andMV were 700 µM and 2,500µM in the MVassay and 55µM and 75 µM for nitrate and Fd in the Fd-assay.The enzyme was inhibited by thiol reagents and metal-chelatingreagents. (Received October 1, 1982; Accepted March 8, 1983) 相似文献
11.
Effects of Nitrate Withdrawal and Resupply on the Assimilation of Nitrate in Leaves of Tomato 总被引:4,自引:0,他引:4
Nitrate reduction in leaves of tomato occurred at the same ratein plants grown in 8.0 mol m3 nitrate as in plants grownin 2.0 mol m3 nitrate, but at a much slower rate in plantsgrown in 0.1 mol m3 nitrate. However, the plants grownin 8.0 mol m3 nitrate had a larger leaf system than theplants grown in 2.0 mol m3 nitrate, and so the totalcapacity to assimilate nitrate was greater in the plants grownin the higher concentration. It was shown that plants grownin 8.0 mol m3 nitrate were better buffered against nitratewithdrawal than plants grown in 2.0 mol m3 nitrate asthe rate of nitrate reduction declined more slowly when plantswere transferred to 0.1 mol m3 nitrate from the higherconcentration than from the lower concentration. Furthermore,leaf expansion continued in the plants transferred from thehigher concentration, whereas it ceased abruptly in the plantstransferred from the lower concentration. It was concluded thatboth continuing expansion and continuing nitrate reduction wereaccompanied, and possibly caused by, a release of nitrate fromstorage pools in the lower part of the stem or the roots. Duringwithdrawal of nitrate the leaves were shown to maintain potentialactivity of the enzyme nitrate reductase although there wasno nitrate to be reduced. When nitrate was resupplied it couldbe reduced very quickly and reduction in the leaves was seento increase within 5 h of resupply. By 3 d after resupply furtherenzyme activity had been induced. Key words: Lycopersicon esculentum Mill, nitrate assimilation, nitrate reductase activity, nitrate withdrawal 相似文献
12.
The changes in nucleic acid and protein accumulation during the batch cultivation of sugarcane cells
Growth and changes in the nucleic acid and protein contentsof sugarcane suspension cultures have been investigated. Theweight of cells in a sugarcane culture increases exponentiallyand then linearly following fresh inoculation. The amount ofRNA and cytoplasmic protein per unit weight of cells in a cultureincreases and then decreases rapidly; that is, no steady stateperiod was found for the accumulation of these nucleic acidsand proteins, not even during a period of exponential cell increase.Changes in DNA content are less pronounced than in RNA content. Without the addition of 2,4-D to a medium containing yeast extract,the weight of cells increases at a faster rate than in a mediumwith 2,4-D. This suppression of weight increase takes placeeven when the concentration of 2,4-D is as low as 0.05 µg/ml.The weight increase of the cells in a medium without 2,4-D,however, is not accompanied by comparable accumulation of RNA.
1 Published with the approval of the Director as Paper No. 371in the Journal Series of the Experiment Station, Hawaiian SugarPlanters' Association, Honolulu, Hawaii; U.S.A. (Received November 1, 1974; ) 相似文献
13.
Spinach (Spinacea oleracea L.) nitrate reductase (NR) is inactivated by phosphorylation on serine-543, followed by binding of the phosphorylated enzyme
to 14-3-3 proteins. We purified one of several chromatographically distinct NRserine-543 kinases from spinach leaf extracts, and established by Edman sequencing of 80 amino acid residues that it is a calcium-dependent
(calmodulin-domain) protein kinase (CDPK), with peptide sequences very similar to Arabidopsis CDPK6 (accession no. U20623; also known as CPK3). The spinach CDPK was recognized by antibodies raised against Arabidopsis CDPK. Nitrate reductase was phosphorylated at serine-543 by bacterially expressed His-tagged CDPK6, and the phosphorylated
NR was inhibited by 14-3-3 proteins. However, the bacterially expressed CDPK6 had a specific activity approx. 200-fold lower
than that of the purified spinach enzyme. The physiological control of NR by CDPK is discussed, and the regulatory properties
of the purified CDPK are considered with reference to current models for reversible intramolecular binding of the calmodulin-like
domain to the autoinhibitory junction of CDPKs.
Received: 12 February 1998 / Accepted: 28 May 1998 相似文献
14.
S-Adenosyl-L-methionine-magnesium-protoporphyrin IX methyltransferase(EC 2.1.1.11
[EC]
) is present in greening barley seedlings associatedwith the particulate fraction. This enzyme was purified 20 foldusing protamine and ammonium sulfate precipitation. The enzymewas active over a wide pH range with highest activity at pH7.5. The Km values for Mg-protoporphyrin IX and S-adenosylmethioninewere 48 and 39 µM, respectively; S-adenosylethionine andS-adenosyihomocysteine were competitive inhibitors with respectto S-adenosylmethionine; hemin inhibition was non-competitivewith respect to Mg-protoporphyrin IX; thiol compounds exhibiteda stimulatory effect on enzyme activity. The properties of theenzyme are discussed and compared with the enzyme from otherorganisms.
1 This research was supported in part by the Utah State AgriculturalExperiment Station.
2 Present address: Department of Chemistry, Boston University,Boston, Massachusetts, U. S. A.
3 Present address: Department of Biochemistry and Microbiology,Faculty of Pharmacy, Comenius University, Bratislava, Czechoslovakia. (Received February 20, 1978; ) 相似文献
15.
This paper describes the first experimental evidence that theinhibition of nitrate reductase by Mg2+ or Ca2+ is related tothe hysteretic properties of the enzyme. The low activity formof nitrate reductase, i.e. the form of nitrate reductase showinghysteretic behaviour, was inhibited 7090% by 5 mM Ca2+or Mg2+. However, no inhibition by Ca2+ or Mg2+ was seen afterthe enzyme was converted to its high activity form by preincubationwith substrates. Addition of thiol compounds or certain aminoacids to the assay mixture also prevented the Mg2+ or Ca2+ inhibition. (Received June 28, 1993; Accepted August 11, 1993) 相似文献
16.
《Plant Science Letters》1979,14(1):85-90
Spinach plants were grown for 8 weeks in sand culture with either complete nutrient containing molybdate (0.05 p.p.m.) or molybdenum-free nutrient containing tungstate (1 p.p.m.). Nitrate reductase and its tungsten analogue were extracted and purified. Nitrate reductase activities (units/kg leaf) decreased during purification from 22.72 to 4.59 (molybdate plants) and from 2.19 to 0.86 (tungstate plants). Cytochrome c reductase activities decreased from 211 to 26 (molybdate plants) and from 539 to 22 (tungstate plants) reflecting the super-induction of the small molecular size enzyme and decreased stability of the analogue. Both nitrate reductase and its analogue contained similar amounts of cytochrome b557kg leaf, which was reduced by NADH and reoxidised by excess of the dehydrogenase electron acceptor dichlorophenolindophenol. Only the haem in the enzyme from molybdate plants was reoxidisable by nitrate. 相似文献
17.
Summary Ten nitrate reductase (NR)-deficient mutants have been characterized for their cross-reactivity against specific barley (Hordeum vulgare L.) nitrate reductase antibodies. The rabbit antibodies raised against the purified barley wild type (cv. Steptoe) enzyme quantitatively inactivate nitrate reductase in crude extracts. All nitrate-grown (induced) mutants show positive precipitin reaction against the antiserum by Ouchterlony double diffusion test and all have the ability to neutralize antisera in a NR protection assay. Under induced growth conditions, mutants Az 12, Az 23, Az 29 and Az 30 which have low NR associated catalytic activities also have the lowest level of antigenicity; mutants Az 13, Az 31, Az 33 and Az 34 have intermediate level of both NR associated catalytic activities and antigenicity, while mutants Az 28 and Az 32 have the highest level of both NR associated catalytic activities and antigenicity. Under noninduced growth conditions, all mutants except Az 12 contain detectable but very low levels of NR antigenicity. These results support the concept that these NR-deficient mutants with various levels of NR associated catalytic activities represent different mutation events at the loci coding the NR structural components.Abbreviations NR
nitrate reductase
- DTT
dithiothreitol
- FAD
flavin adenine dinucleotide
- BSA
bovine serum albumin
- NRCRM
nitrate reductase cross-reacting materials
Scientific Paper No. 5765. College of Agriculture Research Center, Washington State University, Pullman, Project Nos. 0233 and 0430. Supported in part by National Science Foundation Grant #PCM7807649, and U.S. Department of Agriculture CRGO Grant #7900536 相似文献
18.
The induction of nitrate reductase by molybdenum or nitratein excised tissues of cauliflower leaf was dependent on temperature;for the range 2? to 12? C, Q10 was about 2; for the range 12?to 22? C, Q10 was greater than 3. Enzyme formation was initiallymost rapid at 32? C but did not continue for as long as it didat 22? or 24? C. Decreased oxygen supply lessened the rate ofenzyme formation. The effects on enzyme formation of a widerange of natural and synthetic antimetabolites were tested withrespect to induction by either nitrate or molybdenum, when introducedat the same time by infiltration. Actidione (cycloheximide),patulin, cycloserine, polymyxin B, L-2-thiolhistidine D-methionine,L-dihydroxyphenylalanine, D,L--methylglutamic acid, sarcosineand 1 ,2-dichloro-4-(p-nitrobenzenesulphonylamido)-5-nitrobenzene(DCDNS) were the most inhibitory compounds tested. Serine stimulatedproduction of enzyme activity; kinetin, benzimidazole, and p-fluorophenylalanine,3--methyltryptophane and the 4- isomer, chloramphenicol, gramicidin,and several thio- andaza- derivatives of purines or pyrimidineswere practically without effect. Differential effects of inhibitorson enzyme formation in response to nitrate or molybdenum wererarely observed, and no deductions regarding the possible sequencein which the substrate and prosthetic metal induce activitycould be inferred from the results. 相似文献
19.
An investigation was made to determine the effects of univalentcations as activators on the formation of nitrate reductaseand nitrite reductase in rice seedlings. K+ functioned moreeffectively as a univalent cation activator than did other univalentcations examined. Substitution of Rb+ for K+ resulted in stimulationof nitrate reductase formation at about half the rate obtainedwith K+. There was no effect on nitrite reductase formation.Na+ could be partially substituted for K+ in the formation ofboth enzymes. NH4+ slightly inhibited formation of the enzymes.In the absence of univalent cations, enzyme formation proceededat a slower rate during the initial 15-hr period, but thereafterproceeded at a higher rate. This delayed formation was not observedin the presence of K+. Results from inhibitor experiments suggestthat K+ stimulates the formation of nitrate reductase and nitritereductase. In conclusion, when nitrate nitrogen is supplied to rice plantsutilization of the nitrogen may be accelerated by increasedformation of enzymes involved in nitrate assimilation in thepresence of K+. (Received February 21, 1969; ) 相似文献
20.
In vivo nitrate reductase activity was measured over the seasonin individual organs of the main tiller of the euploid and fourditelosomics of the wheat variety Chinese Spring. A generalbiphasic profile of leaf activity was obtained in vivo and invitro, the peaks corresponding to emergence of leaf 7 (the pre-flagleaf) and the ear. A wide range of seedling nitrate reductaseactivity was exhibited by these stocks and a significant positivecorrelation was obtained between seedling activity and the dailymean of activity integrated over the season. Seasonal euploid-ditelosomicdifferences in nitrate reductase activity reflected differencesover the vegetative stage, but no significant inter-stock differencesin activity were found over the reproductive stage. The highseasonal nitrate reductase activity of ditelo-7BL and 7BSappeared to be due largely to high mean activities of individualleaves, while that of ditelo-4BL depended on the longer durationof active tissue than was exhibited by the euploid. Significantactivity was assayed in non-leaf organs, especially the rachisand awns, and constituted an important proportion of the totaltiller activity late in the season. 相似文献