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1.
Subunits (, ß, ) and mixtures of subunits ( ß, , ß , ß ) were isolated without denaturationfrom a chloroform extract of chloroplast coupling factor 1 (CF1)from maize (Zea mays var. Ushiku 5-4) and from spinach by fastprotein liquid chromatography (FPLC), on an anion-exchange columnof Mono-Q in the presence of n-octylglucoside (OG) and on achromatofocusing column of Mono-P. The ß -subunitcomplex (CF1 ß ) was the minimum unit required forATPase activity, as was confirmed by the reconstituted complexof ß and subunits. An subunit isolated from maizeinhibited the ATPase activity of CF1 ß from bothmaize and spinach. CF1 ß was found to contain anOG-dependent Mg2+-ATPase. The ATPase activity of CF1 ß required divalent cations, such as Mg2+ or Mn2+, for its expressionin the presence of OG; its optimum pH was 8.0 and it was markedlyinhibited by NaN3. The enzyme hydrolyzed ATP in prefernece toGTP but not CTP, UTP, ADP, AMP or pNPP. Lineweaver-Burk plotsof its activity were curvilinear in the range of 0.60.7mM ATP.Mg2+.
1Present address: Department of Biology, School of Education,Waseda University, Shinjuku-ku, Tokyo, 160 Japan. (Received February 15, 1989; Accepted April 20, 1989) 相似文献
2.
A Ras-related NTP-binding protein was partially purified froma membrane fraction derived from the mycelia of Neurospora crassa.[-32P]ATP and [-32P]GTP were incubated with mem brane and solublefractions which were then irradiated with UV light to inducecrosslinking of tightly bound nucleotides. After SDS-polyacrylamidegel electrophoresis, blotting onto a nitrocellulose filter andautoradiography it was apparent that most of the proteins thatbound [-32P]-GTP also bound [-32P]ATP. Pretreatment of the membranefraction with Ras-specific antibody effectively blocked thebinding of [-32P]ATP and [-32P]GTP to several ATP-GTP-bindingproteins. The band of a protein with a molecular weight of 26kDa on the SDS-polyacrylamide gel cross-reacted strongly withthe Ras-specific antibody. The protein was extracted from thegel and further purified by repeated gel electrophoresis. Thepurified protein bound [-32P]ATP, [-32P]-GTP, [-32P]CTP and[-32P]UTP at 1.6x10 M and was autophosphorylated in thepresence of [-32P]ATP and [-32P]GTP at 1.7x10 M. Pretreatmentof the protein with Ras-specific antibody partially blockedthe autophosphorylation in the presence of these nucleotides.The binding of [-32P]ATP to the NTP-binding protein was blockedby addition of ATP at 104103 M. ATP ata concentration of 104 M prevented the binding of [-32P]to a greater extent than did GTP at the same concentration.Binding of [-32P]CTP and [-32P]UTP to the protein was also observed. (Received October 7, 1991; Accepted July 14, 1992) 相似文献
3.
A b-type cytochrome having an -band at 560 nm was isolated fromspinach leaves (Spinacia oleracea). A method is described forpreparing this cytochrome, cytochrome b-560 (spinach), in apurified state. The cytochrome has, in its reduced state, absorption bands at560 nm (), 530 nm (ß) and 427 nm (); and in the oxidizedstate at 562 nm (), 529 nm (ß) and 417 nm (). Thepyridine ferro-haemochrome prepared from cytochrome b-560 hadan -band at 556.5 nm, indicating the protohaem-nature of theprosthetic group. The cytochrome has an oxidation-reduction potential (E'0) of+0.13V at pH 7.0, as measured using the ferri-ferro oxalate system. The cytochrome is rapidly reduced on illumination with red orfar-red light in the presence of spinach chloroplasts and isoxidized at a slower rate in the dark. This photoreduction isinhibited by 1x106 M 3-(3,4-dichlorophenyl)-1,1-dimethylurea(DCMU). The molecular weight of the cytochrome is 30,000 asestimated by the dextran gel filtration method. (Received December 3, 1971; ) 相似文献
4.
Nucleoside triphosphate(NTP)-binding proteins were detectedin the crude extract of mycelia of Neurospora crassa, whichwas treated with 1% Lubrol PX and fractionated by gel filtration.Protein fractions showing the capacity to bind [35S]ATPS or[35S]GTPS were designated as AGN1 to 6. The binding of [35S]ATPSor [35S]GTPS was prevented in the presence of 0.1 mM ATP orGTP except that in fractions AGN1 and 2, the presence of GTPstimulated the binding of [35S] ATPS to ATP(NTP)-binding proteins.ATP or GTP was 1 to 2 orders of magnitude more effective thanCTP or UTP in preventing the binding of [35S]GTPS in AGN1, 2and 5. Among these fractions AGN1, 2, 5 and 6 showed activityto hydrolyze 1 nM [32P]ATP or [32P]GTP. NTP-bindingproteins bound with [35S]ATPS or [35S]GTPS had lower apparentmolecular weights than the same proteins without bound nucleotide.Proteins bound with [35S]ATPS or [35S]GTPS and those [32P]ADP-ribosylatedby endogenous ADP-ribosyl transferase in each fraction wereanalyzed by SDS-PAGE. About 20 species of ATP or ATP-GTP-bindingproteins were detected, several of which were ADP-ribosylated.The binding of [35S]ATPS or [35S]GTPS to NTP-binding proteinswas confirmed by the comparison of non-boiled and boiled samplesimmediately before loading to SDS-PAGE. ATP, GTP, CTP or UTPat the concentration of 0.1 mM effectively removed [33S]ATPSor [35S]GTPS bound to NTP-binding proteins. (Received December 10, 1990; Accepted April 18, 1991) 相似文献
5.
Effects of l, N6-ethenoadenylates (e-adenylates) were testedon phosphorylation, and electron transport under phosphorylation,arsenylation and quasi-arsenylation (stimulation of electrontransport in the presence of ATP, AMP and arsenate) conditionsin isolated spinach chloroplasts. -ATP as well as ATP partially inhibited ferricyanide reductionthrough binding to the chloroplast coupling factor 1 with anapparent dissociation constant (KDapp) of around 5µM,which was remarkably larger than that for ATP (ca. 2µM).e-ATP at below 500 µM had no effect on phosphorylationbut inhibited quasi-arsenylation in competition with ATP withan apparent inhibition constant (K1app) of around 60 µM. -ADP as well as ADP partially inhibited ferricyanide reductionwith a KDapp value close to that for -ATP. -ADP was phosphorylated(the apparent Michaelis constant, Kmapp=80µM) accompanyingstimulation of ferricyanide reduction to the magnitude predicted(P/e=l). -ADP-arsenylation was also detected by stimulationof ferricyanide reduction. -AMP alone caused little inhibition of ferricyanide reductionas AMP, but competitively depressed the electron transport inhibitionby ADP and ATP with a K1app value of around 200 µM. -AMPwas not effective for ADP phosphorylation but inhibited stimulationdue to quasi-arsenylation coupling in competition with AMP K1app=150µM Among the possible combinations of adenylates and -adenylatesfor quasiarsenylation, only [ATP+AMP] could couple with theenergy transduction mechanism. Based on the specificity of binding sites to adenylates and-adenylates, an attempt was made to distinguish at least four(two pairs) kinds of binding sites (at least six sites in toto)on the chloroplast coupling factor 1 for photosynthetic energytransduction. When one pair of sites is occupied by the designatedadenylates or -adenylates (allosteric effectors), the couplingfactor is thought to be in a conformation for coupling withthe energy transduction mechanism in the presence of phosphateor arsenate.
1Presented to the 1st Symposium of Japan Bioenergetics Group,December 19, 1975, Osaka. (Received February 17, 1976; ) 相似文献
6.
Yanagishima N.; Yoshida K.; Hamada K.; Hagiya M.; Kawanabe Y.; Sakurai A.; Tamura S. 《Plant & cell physiology》1976,17(3):439-450
The sexual agglutinability of haploid cells of heterothallicSaccharomyces cerevisiae was repressed when they were culturedin the absence of easily fermentable sugars, such as glucoseand mannose. The repression was reversed by the action of hormone-likesubstances of the opposite mating types. The substance producedby mating type cells was identical to subtsance-I which isknown to induce sexual agglutinability of inducible matingtype cells. The mating type cells produce a new hormone-likesubstance which induces or enhances sexual agglutinability of mating type cells. A crude fraction of the mating type-specific substance ( substance-I)was obtained by passing the culture filtrate of mating typecells through Amberlite CG-50 (H+ form), followed by elutionwith 1.5 M ammonia.
2 On leave from Osaka City University. (Received December 25, 1975; ) 相似文献
7.
Enzymatic Degradation of Chlorophyll in Chenopodium album 总被引:3,自引:0,他引:3
The breakdown of chlorophyll (Chi) in crude extracts of Chenopodiumalbum (white goose foot) in the dark was examined. Derivativesof pheophorbide were formed when Chi or chlorophyllide wasincubated with depigmented crude extracts. The formation ofpheophorbide was completely prevented by heat treatment of extracts,indicating that the reaction was enzymatic, and the presenceof a Mg-releasing enzyme, the so called Mg-dechelatase, waspostulated. This hypothesis was strongly supported by the observationthat the formation of pheophorbide was inhibited by 51% by 10mM MgCl2. Analysis by high-performance thin-layer chromatography(HPTLC) and liquid chromatography (HPLC) showed that the appearanceof chlorophyllide , pheophorbide 132-hydroxychlorophyllide and pyropheophorbide was accompanied by a concomitant decreasein levels of Chi The formation of 132-hydroxychloro-phyllide was not clearly an enzymatic reaction and requires furtherexamination. It appears that Chl is degraded in a crude extractof C. album via the following enzymatically catalyzed reactions
(Received September 10, 1990; Accepted November 15, 1990) 相似文献
8.
Uptake of Thallium, a Toxic Heavy-Metal, in the Cyanobacterium Synechococcus R-2 (Anacystis nidulans, S. Leopoliensis) PCC 7942 总被引:1,自引:0,他引:1
Uptake of the toxic heavy-metal, thallium, was studied in thecyanobacterium Synechococcus R-2 (PCC 7942) using clinicallyavailable 201Tl +. Thallium was found to distribute across theplasmalemma passively, and so the accumulation ratio of theion ([Tl+]i/[Tl+]o) could be used to calculate the apparentmembrane potential (i,o) of the cells (ETI+i,o = i,o).The permeability of the plasmalemma to TI+ (PTI+ 1 to 5 nms1)is higher than that of K+. Valinomycin does not increase thepermeability of TI+. Transient changes in the i,o of cells,because of electrogenic transport of ions, could be detectedfrom its effects upon the uptake rate of TI+. HCO3 hyperpolarizedSynechococcus cells, whereas NH+4, CH3NH+, and K+ led to depolarization.The use of TI+ as a reporter of i,o has some inherent limitations.Tl+ is toxic at very low concentrations (inhibitory effectsare apparent after about 6 h at concentrations as low as 1 mmolm3). The rate of equilibration is slow (t1/25 to 20 min).Equilibration of TI+ takes about 2 h, which limits its valueas a membrane potential probe. Large amounts of TI+ bind tothe surface of the cells making the method impracticable formeasuring accumulation ratios of less than about 10 (i,o)values smaller than about 60 mV). Cultures continuouslyexposed to Tl+ (10 mmol m3) eventually become TI+ resistantby actively extruding TI+ (µTI+i,o= 3±0.2kJ mol1) and so thallium cannot be used as a i,oprobe in such cells. (Received October 28, 1997; Accepted August 31, 1998) 相似文献
9.
Yoneyama Tadakatsu; Fujita Kounosuke; Yoshida Tomio; Matsumoto Tetsuo; Kambayashi Iwao; Yazaki Jinya 《Plant & cell physiology》1986,27(5):791-799
Sixteen legumes were grown in N-free media so that N was suppliedentirely by symbiotic N2 fixation. The plant tissues were analyzedfor natural 15N abundance (expressed as 15N per mil relativeto air N2) with a ratio mass spectrometer. The nodules of desmodium,centro, siratro, soybean and winged bean showed high enrichmentin 15N (+9), while red clover showed slight enrichment (+2).The nodules of 9 other forage legumes (Townsville stylo, whiteclover, alsike clover, common vetch, Chinese milk vetch, senna,alfalfa, ladino clover, and hairy vetch) showed little enrichmentin 15N. In all the legumes investigated, particularly in the ureide-transportingplants such as desmodium, centro, siratro, soybean, winged beanand field bean, the 15N value of the shoots was negative (3.2).The 15N value of the shoots in winged bean and field bean variedby about 1 depending on the Rhizobium strains used. The isotopicmass balance of 13 legumes indicated that isotopic fractionationoccurs during N2 fixation by the legume-rhizobia symbiosis witha preference for 14N over 15N, resulting in a 15N value of 0.2to 2 in the whole plant. The results indicate that 15N/14N isotopic discrimination witha preference for the lighter atom may occur in both N2 fixationand export of fixed N from nodules.
1Present address: Department of Soils and Fertilizers, NationalAgriculture Research Center, Kannondai, Tsukuba, Ibaraki 305,Japan. (Received October 8, 1985; Accepted April 7, 1986) 相似文献
10.
Various Cucurbita seed globulins showed patterns similar toone another on SDS-gel electrophoresis, and ß bandsfor unreduced globulins and , ', and ' bands for reduced ones.On gel electrophoresis in 6 M urea, reduced globulin gave twoacidic and two basic bands. These corresponded to and ' chainsand 1 and 2 chains, respectively, identified by two-dimensionalurea-SDS gel electrophoresis. The compositions of the and ßsubunits were proposed. (Received September 8, 1977; ) 相似文献
11.
1. Localization of carbon in caffeine molecule biosynthesizedfrom the N-methyl carbon of -glutamylmethylamide in tea plantswas observed. 14C-Caffeine produced from 14C--glutamylmethylamidewas isolated and degraded. Approximately 2655% of the14C was observed in the three methyl carbons in caffeine, withonly 23% at the C-2 carbon, 37% at the C-8 carbonposition. The amount of 14C at the C-4, C-5 and C-6 positionswas calculated from the results obtained. 2. The role of the N-methyl carbon of -glutamylmethylamide inthe formation of RNA in tea plants was examined. Incorporationof the N-methyl-14C of 14C--glutamylmethylamide into AMP andGMP in RNA was found. These facts indicate that in tea plants, -glutamylmethylamideis metabolized and most of its N-methyl carbon is utilized asa precursor for caffeine formation and little, if any, as aprecursor for nucleic acid formation.
1 Present address: Department of Agricultural Chemistry, ShizuokaUniversity, Iwata, Shizuoka 438, Japan. (Received February 2, 1972; ) 相似文献
12.
The luciferin-luciferase method was used to determine ATP extractedfrom darkmaintained and light-exposed samples of the green algaChlorella pyrenoidosa and of the blue-green alga Anacystis nidulans.A few measurements on Synechococcus lividus (a bluegreen thermophile,clone 65?C) are also reported.
- The light-minus-dark ATP levels (ATP) from aerobic cells ofChlorella and Anacystis were negative; however, ATP from Synechococcuswas positive. Large positive ATP was obtained in regularly grown(RG: moderate light) Chlorella treated with oligomycin; darklevels were reduced, light levels remained essentially unaffected.In high-light exposed (HLE) Chlorella, oligomycin reduced bothlight and dark ATP levels, but positive ATP was still obtained.However, in Anacystis, which has a different organization ofthylakoid membrane, oligomycin severely reduced both the lightand the dark ATP levels and the ATP remained negative.
- Theoligomycin (12 µM) treated Chlorella and the untreatedAnacystis and Synechococcus show the presence of cyclic photophosphorylationunder conditions in which the non-cyclic electron flow fromphotosystem II to photosystem I is blocked by 10 µM 3-(3,4-dichlorophenyl)-l,l-dimethylurea(DCMU), or not allowed to operate by the absence of CO2. Cyclicphotophosphorylation ranged from 1030% of the maximumATP in RG, to 4050% in HLE Chlorella. In RG Chlorella,cyclic and non-cyclic (in the absence of DCMU) photophosphorylation(ATP) saturate at about 103 ergs cm2 sec1 and104 ergs cm2 sec1 and 104 ergs cm2 sec1red (>640 nm) light, respectively; a lag was observed inthe light curve.
- In Chlorella, the addition of the photosystemI electron acceptormethyl viologen (MV; 1 mM) increased ATPby twofold. Furtheraddition of DCMU (25 µm) reduced thisto the level observedwith DCMU alone. If 1 mM reduced dichlorophenolindophenol orphenazine methosulphate (DCPIPH2 or PMSH2, respectively)wasadded along with DCMU, the ATP level was 3040% ofthecontrol. Further addition of MV increased the JATP to be7080%of that of the control. These and other resultsconfirm thepresence of both non-cyclic and cyclic photophosphorylationin vivo, the former predominating in Chlorella, and the latterin Anacystis and Synechococcus.
13.
In Vigna mungo cotyledons, the -amylase activity increased markedlyduring germination at 27°C in the dark, while the activityof other amylases was very low. The -amylase was purified from4-day-old cotyledons by affinity chromatography on epoxyactivatedSepharose 6B substituted with rß-cyclodextrin andby column chromatography on Bio-Gel P-200. Gel filtration andpolyacrylamide gel electrophoresis showed that the enzyme existsmostly as a monomer (43,000 daltons), but partially aggregatesto form dimer, trimer and further multimers. Ca2+ protectedthe -amylase against heat inactivation. Incubation of the enzymewith 5 mM EDTA or dialysis against 10 mM EDTA resulted in a5090% loss of activity. The inactivation was partiallyreversed by the addition of Ca2+. Other properties, such asthe amino acid composition, Km value, pH optimum and activationenergy were similar to those of other plant -amylases. (Received May 6, 1981; Accepted June 22, 1981) 相似文献
14.
15.
A gamete recognition mechanism in Antithamnion sparsum Tokidais proposed based on experiments using various lectins and carbohydrates.Spermatial binding to trichogynes is inhibited by pre-incubationof spermatia with concanavalin A (ConA) and/or L-fucose, whiletrichogyne receptors are blocked by the complementary carbohydrate-methyl D-mannose and/or the lectin Ulex europaeus agglutinin(UeA1). Binding inhibition (4050%) was observed with1050 mM carbohydrates and 2550 µg ml-1 lectins.The inhibitory effects of ConA and UeA1 is partially reversed(to 8090% of controls) by addition of -methyl D-mannoseand L-fucose, respectively. Lectin binding to spermatial surfaceswas visualized by Fluorescein isothiocyanate (FITC) conjugatedConA, whereas carbohydrate receptors along the trichogyne andspermatium were localized with -mannosylated-FITC-albumin andL-fucosylated-FITC-albumin, respectively. These results suggestthat gamete recognition in Antithamnion sparsum is mediatedby a double-docking recognition system consisting of spermatiapossessing surface L-fucose receptors and -methyl D-mannosemoiety, and trichogynes possessing the complementary receptors. (Received December 5, 1995; Accepted April 22, 1996) 相似文献
16.
-Aminolevulinic acid dehydratase (5-aminolevulinate hydro-lyase,EC 4.2.1.24
[EC]
) was purified from greening radish cotyledons. Thefinal product was homogeneous on polyacrylamide disc gel electrophoresisand had a molecular weight, estimated by gel filtration, of282,000 daltons. The enzyme seems to require magnesium ion aswell as sulfhydryl compounds for maximum activity. EDTA anda low concentration of zinc ion markedly inhibited the activity.The optimum pH was 8.0; the Km value for -aminolevulinic acidwas 3.85?104M. Levulinic acid was a competitive inhibitorof the enzyme, with a Ki of 2.14?104M. These propertieswere compared with those of microorganism and animal -aminolevulinicacid dehydratases. (Received November 22, 1976; ) 相似文献
17.
-Aminolevulinic acid dehydratase (-aminolevulinic acid hydrolyaseEC 4.2.1.24
[EC]
) which catalyzes the formation ofporphobilinogenfrom two molecules of -aminolevulinic acid (ALA) was purifiedfrom Chlorella regularis 737-fold by acetone and ammonium sulfatefractionations, DEAE-cellulose column chromatography, and SephadexG-200 gel filtration. The enzyme had an optimum pH of 8.5 inTris-HCl buffer and required either Mg2+ or Mn2+ for its maximumactivity. The Km values for Mg2+, Mn2+ and ALA were 15 µM,10µM, and 0.5 mM, respectively. The enzyme was not activatedby thiol compounds, but was inhibited by p-chloromercuribenzoate.The molecular weight estimated by gel filtration was 316,000and the isoelectric point was 5.25. (Received October 18, 1978; ) 相似文献
18.
Cell walls of 4-day old rice seedlings were extracted successivelywith ammonium oxalate-oxalic acid, 4% KOH and 24% KOH. A rß-D-glucanpreparation and a xyloglucan preparation were isolated fromthe 4% KOH extract and 24% KOH extract, respectively. Methylationanalysis and enzymic degradation studies of the polysaccharidesshowed that the former was built up predominantly of repeating-oligosaccharideunits of 3-O-rß-cellobiosyl-D-glucose and 3-O-rß-cellotriosyl-D-glucosein a molar ratio of 2.6 : 1.0, and the latter was of repeating-oligosaccharideunits of -D-xylosyl-(16)-rß-D-glucosyl-(14)-[-D-xylosyl-(16)]-rß-D-glucosyl-(14)-D-glucose,-D-xylosyl-(16)-rß-D-glucosyl-(14)-D-glucose and cellobiose.
1 Present address: Department of Botany, Iowa State University,Ames, Iowa 50011, U.S.A. (Received August 29, 1981; Accepted January 12, 1982) 相似文献
19.
Two proteolytic activities I and II involved in the globulindegradation were detected in pumpkin seeds. Activity I, hydrolyzing and ß subunits of the globulin to form Fß,was found in both dry seeds and cycloheximide-treated cotyledons,and decreased during germination. Activity II, hydrolyzing Fßto produce small peptides and amino acids, was not observedin dry seeds but found in cycloheximide-treated cotyledons,increased up to 4 days, and gradually decreased during germination. Activity I gave limited hydrolytic products from the globulinand the chain, but not from Fß, the chain and some animal proteins. It was inhibitedby EDTA. On the other hand, activity II hydrolyzed Fßand the chain faster than the globulin, the chain and some animal proteins. It was inhibitedby EDTA and p-chloromer-curibenzoate, and activated by ß-mercaptoethanol,dithiothreitol and CoCl2. Optimum pH's were at about 6.8 andat 6.0 to 6.8 for activities I and II, respectively. The degradation process of the globulin can be divided intotwo steps: the first step is the conversion of globulin to Fßand the second step, Fß to small peptides and aminoacids. (Received November 9, 1979; ) 相似文献
20.
Tea seedlings were treated with 14C-methylamine to cause synthesisof 14C--glutamylmethylamide (N-methyl-14C). The metabolic conversionof -glutamylmethylamide was studied by tracing 14C. 14C--Glutamylmethylamide (N-methyl-14C) translocated from rootsand cotyledons to shoots of tea seedlings, was converted almostentirely into caffeine. Conversion was greater in light-exposedsamples. For those grown in the dark, the converted amount didnot correspond to the total caffeine produced. More 14C--glutamylmethylamidewas present in stems than in leaves, but with 14C-caffeine,the opposite was found. When 14C--glutamylmethylamide or 14C-methylamine was appliedto leaf disks, 14C-caffeine was biosynthesized from both substances.
1 Present address: Department of Agricultural Chemistry, ShizuokaUniversity, Iwata, Shizuoka 438, Japan. (Received September 25, 1971; ) 相似文献