Characterization of membrane-bound Mg++-activated ATPase isolated from the lower epidermis of tobacco leaves

Membrane-bound Mg⁺⁺-activated ATPase was separated from thelower epidermis of tobacco leaves (Nicotiand tabacum L. SamsunNN) on stepwise sucrose density gradient centrifugation. Membrane-bound epidermal ATPase was localized in the interfaceof densities in sucrose of 1.12 to 1.16 in the sedimentary fractionbetween 1,500?g to 10,000?g from the homogenate of the lowerepidermis. The epidermal ATPase activity was activated by divalentcations (Mg⁺⁺>Mn⁺⁺Co⁺⁺>Fe⁺⁺>Zn⁺⁺>Ca⁺⁺) and furtherstimulated by KCl by ca. 20%. The pH optimum for Mg⁺⁺-activationof the epidermal ATPase was ca. 6.0. The enzyme hydrolyzed ATPmore rapidly than other nucleoside triphosphates. The optimumtemperature for activation of the epidermal ATPase activitywas ca. 40?C. 50% of the epidermal ATPase activity was lostin 18 min at 55?C and in 2.5 days at 2.5?C. The apparent Kmvalue of the epidermal ATPase was 4.7?10^–4 M and Vmaxwas 65.4 nmoles Pi/mg protein/min. The epidermal ATPase wasstrongly inhibited by N, N'-dicyclohexylcarbodiimide (DCCD)in vitro whereas oligomycin, carbonyl cyanide m-chlorophenylhydrazone(CCGP), indoleacetic acid (IAA) and abscisic acid (ABA) wereinsensitive to the epidermal ATPase activity. (Received May 23, 1978; )     

Studies on NADP-isocitrate dehydrogenase from maturing castor bean seeds

NADP-specific isocitrate dehydrogenase from the soluble fractionof maturing castor bean endosperm was partially purified (approximately180-fold) and some of its enzymatic properties were studied.Mg⁺⁺, Mn⁺⁺, Cd⁺⁺, Ba⁺⁺, Co⁺⁺, Zn⁺⁺, and Sr⁺⁺ were activatorsof the enzyme reaction at a concentration of 6.7x10 M. The optimumpH of this enzyme was about 8.5. The enzyme was stable in thenarrow range from pH 7.0 to pH 8.0. Km values for isocitrateand NADP at pH 8.5 were 3.5x10^–6 M and 3.6x10^–6M, respectively. Enzyme stability was not affected by NaCl concentrationand enzyme reaction was inhibited at 5x10^–6 M PCMB (80%inhibition). It is suggested that the condensation product ofglyoxylate and oxalacetate also inhibits the reaction. NADP-IDHin the crude extract from maturing castor bean endosperm washeat-stable but the dialyzed enzyme preparation and the partiallypurified enzyme were labile against heat treatment at 57°C.When Mg⁺⁺ was added to the partially purified enzyme in thepresence of isocitrate or NADP, the enzyme was stabilized againstheat treatment. Mn⁺⁺, Ca⁺⁺, Co⁺⁺, Sr⁺⁺ or Ba⁺⁺ could be substitutedfor Mg⁺⁺. Addition of only one of the factors, Mg⁺⁺, isocitrateor NADP, had no effect on the heat stability. Moreover, a combinationof isocitrate and NADP did not establish stabilization. A divalentcation plays a central role, while adenine nucleotide, especiallyATP, may have an important part in stabilization. (Received August 14, 1972; )     

Ribulose-1, 5-diphosphate carboxylase of Chromatium strain D

RuDP carboxylase isolated from autotrophically grown cells ofphotosynthetic sulfur bacterium, Chromatium strain D, was partiallypurified by (NH₄)₂SO₄ precipitation and Sephadex G-200 gel filtration.The molecular size of the bacterial RuDP carboxylase was foundto be large, analogous to that of the plant enzyme, in agreementwith results of previous workers. Sucrose density gradient centrifugationshowed the S_rel to be approximately 18; the omission of Mg⁺⁺caused no dissociation of the enzyme molecule in its subunits.Chromatium RuDP carboxylase showed similarities to the plantenzyme in some of its kinetic properties; (a) a shift of pHoptimum to the neutral side from the alkaline side on the additionof Mg⁺⁺, (b) deviation of the substrate concentration (NaHCO₃)-activityrelationship from the MICHAELIS formula and (c) a marked stimulativeeffect of Mg⁺⁺. A unique sigmoidal saturation curve of the enzymeto RuDP, which had been detected in Rhodospirillum rubrum andRhodopseudomonas spheroides RuDP carboxylase in the absenceof Mg⁺⁺, was not found. Another characteristic feature of ChromatiumRuDP carboxylase is its partial immunological response to therabbit anti-spinach RuDP carboxylase serum as detected by theinhibition of the carboxylation reaction due to the antibody-antigenreaction. ¹Part X, Structure and Function of Chloroplast Proteins. Supportedin part by research grants from the Ministry of Education ofJapan (No. 8719) and USPHS (AM-10792-03) (Received July 4, 1969; )     

Effects of Cytokinin and Several Inorganic Cations on the Polyamine Content of Lettuce Cotyledons

Kinetin (4.7 x 10^–5 M) and 6-benzyladenine (2.22 x 10^–5M) were found to increase ca. 2-fold the putrescine contentin cotyledons of lettuce (Lactuca sativa L.) seedlings grownfor 3 days under fluorescent light. On the other hand, severalinorganic ions (K⁺, Na⁺, Ga⁺⁺, Mg⁺⁺) at a concentration of 3x 10^–2 M reduced the putrescine content. The combinationof kinetin with one of several inorganic ions at the same levelmarkedly increased the spermine content, but the putrescinecontent decreased; calcium and magnesium ions were less effective.The physiological significance of these findings is discussed. (Received July 4, 1982; Accepted November 6, 1982)     

Induction of Solute Release from Nicotiana tabacum Tissue 10

For determination of the effects of polymyxin B, polymyxin E,or ethylenediamine tetra-acetic acid (EDTA) on plant cell membranes,the rates at which three solutes, K⁺, P₁, and sugar, leakedfrom treated tissue culture cell suspensions of Nicotiana tabacumwere measured. The kinetics of leakage from cells treated witheither of the polymyxins was biphasic, whereas kinetics forcells treated with EDTA was monophasic. Only K⁺ leaked frompolymyxin-treated cells during the first phase, and all threesolutes leaked during the second phase. The slower first phaseis interpreted as leakage of K⁺ from the Donnan free space andcytoplasm, and the faster second phase as the leakage of solutesfrom the vacuole. The monophasic kinetics of EDTA treatmentindicated that solutes were leaking simultaneously from cytoplasmand vacuole. Of the divalent cations tested, only Ca⁺⁺ and Mn⁺⁺counteracted the effects of polymyxin and EDTA. Ca⁺⁺ even restoredP¹ and sugar uptake. Addition of Mg⁺⁺ or Sr⁺⁺ to polymyxin-treatedcells did not stop solute leakage but actually enhanced theleakage rates. A model is presented that suggests that polymyxinor EDTA induces solute leakage by forming pores in plant cellmembranes. The effects of divalent cations on membranes oncethe pores are formed are also discussed. Key words: Polymyxin, EDTA, Nicotiana tabacum, Solute leakage     

Effect of Root Zone Temperature on the Mineral Composition of Xylem Sap and Plasma Membrane K+-Mg++-ATPase Activity of Grafted-Cucumber and -Figleaf Gourd Root Systems

Cucumber (Cucumis sativus L.) seedlings were grafted onto cucumber-(CG) or figleaf gourd- (FG, Cucurbita ficifolia Bouché)seedlings in order to determine the effect of solution temperature(12, 22, and 32°C) on the mineral composition of xylem sapand the plasma membrane K⁺-Mg⁺⁺-ATPase activities of the roots.Low solution temperature (12°C) lowered the concentrationof NO^–₃ and H₂PO^–₄ in xylem sap of CG plants butnot of FG plants. Concentrations of K⁺, Ca⁺⁺ and Mg⁺⁺ in xylemsap were less affected than anions by solution temperature.The plasma membrane of FG plants grown in 12°C solutiontemperature showed the highest K⁺- Mg⁺⁺-ATPase activity at allATP concentrations up to 3 mM and at low reaction temperatureup to 12°C, indicating resistance of figleaf gourd to lowroot temperature. (Received December 27, 1994; Accepted March 10, 1995)     

Interrelationships between Potassium, Calcium, and Magnesium Nutrition of Zea mays L.

The uptake of potassium, calcium, and magnesium ions by maizeand the interrelationships among the cations have been investigatedat 48 K: Ca: Mg ratios in culture solutions. Calcium was foundto stimulate K⁺ and Mg⁺⁺ uptake at certain cation ratios butinhibit it at others. Potassium did the same for Ca⁺⁺ uptake,and Mg⁺⁺ for Ca⁺⁺ and K⁺. The uptake of Mg⁺⁺ was generally enhancedby K⁺. The sum of the cations in the plants expressed in meqwas fairly constant for treatments of the same K⁺ concentrationat the low to moderate levels of K⁺, but at considerably higher(> 24 meq l^–1) K⁺ levels the constancy was not dependenton K⁺ concentration. Potassium depressed, but Mg⁺⁺ stimulatedphosphorus accumulation. Calcium stimulated phosphate absorptionat certain cation ratios but had no effect at others. The plantyield increased with increasing K⁺ up to 24 meq l^–1 ofK⁺ after which the yield tended to fall with further increasein K⁺. The yield was also increased by Ca⁺⁺. Magnesium increasedthe yield at certain cation ratios and either depressed it orwas without effect at others.     

Application of an Electrokinetic Formulation to the Study of the Effect of Alkaline-earth Cations on the Absorption of K+-ions by Lemna minor

A double effect of Ca⁺⁺ is observed on K⁺-absorption by Lemnaminor: activation through coupled (probably metabolic) processes,and inhibition through a perturbation of the structure of thecatalytical sites of K⁺-absorption. In contrast, Mg⁺⁺, whichalso exhibits a structural inhibitory effect, has not apparentlya metabolic actrvatory one. It seems that diffusive processescould account partly for the second phase of the dual-phasecurves of absorption.     

ATP-ASE ACTIVITY IN ISOLATED FLAGELLA OF CHLAMYDOMONAS SNOWIAE

ATPase in isolated glycerinated flagella of Chlamydomonas snowiaeis shown to be inhibited by high ATP concentrations, by DNPat pH 6.0 and by salyrgan. The enzyme is activated by Mg⁺⁺ andthe activation is reversed by Ca⁺⁺. A number of other factorswhich can affect the phototactic behaviour of the cells do notaffect ATPase activity, indicating that the response of directionalmotility is under separate control in the intact cell. ¹ This work is based on part of an M. Sc. thesis of I.C.-K.     

Metal Interrelationships in Plant Nutrition: 2. THE RELATION OF METAL TOXICITY, MOLYBDENUM AND NITROGEN SOURCE TO CHLOROPHYLL AND MAGNESIUM CONTENT OF BEET IN SAND CULTURE

Sugar beet and spinach beet were grown in sand culture withdifferent sources of nitrogen or iron and different levels ofmolybdenum and heavy metals. Chlorophyll and some magnesiumfractions were determined. Extra molybdenum accentuated chlorosis caused by excess of Mn⁺⁺,Cr⁺⁺⁺, Zn⁺⁺, and Cu⁺⁺ and decreased it in the presence of CrO₄^––with nitrate. In the presence of these ions urea reduced chlorophyll in youngand increased it in old leaves of sugar beet. Effects of ureaand molybdenum were additive and independent. Ammonium sulphatecaused increased chlorosis of spinach beet with Mn⁺⁺ and Zn⁺⁺. The existence of an acetone-soluble magnesium fraction otherthan chlorophyll was shown. Excess Cu⁺⁺ and Zn⁺⁺ decreased totalmagnesium; Mn⁺⁺ did not. Mn⁺⁺ and Zn⁺⁺ reduced chlorophyll,increased the non-chlorophyll acetone-soluble magnesium, butreduced other fractions. Ferrous iron ipcreased total magnesiumcontent and also increased the acetone-soluble fractions inthe presence of Mn⁺⁺ and Zn⁺⁺. Extra molybdenum decreased theacetone-soluble magnesium fractions but did not affect the totalmagnesium content. Significant interactions between Mo, N, Fe, and heavy metalswere also disclosed.     

Some properties of shikimate: NADP oxidoreductase produced in sweet potato root tissue after slicing

The activity of shikimate: NADP oxidoreductase [EC 1. 1. 1.25] in sweet potato root tissue increased soon after slicing.Enzyme preparations obtained from both sliced tissue and fromfresh tissue probably contained a single enzyme component, andthey showed identical chromatographical behaviour. K_m values of the enzyme for NADP and shikimate were 1.0x10^–4Mand 1.3 x 10^–3M, respectively. Enzyme activity was potentlyinhibited by SH-inhibitors such as p-chloromercuribenzoate andoxidized glutathione. Enzyme activity was affected neither by mononucleotides suchas ATP, ADP and AMP, divalent cations, Mg⁺⁺, Ca⁺⁺ and Mn⁺⁺,nor by metabolites such as tryptophan, phenylalanine, tyrosineand t-cinnamic acid which are involved in aromatic compoundsyntheses. The enzyme rapidly lost its activity. This inactivation reactionshowed a time course consisting of two steps of the first-orderreaction. The inactivated enzyme preparation was not reactivatedby thiol compounds such as cysteine, 2-mercaptoethanol and glutathione,although these reagents, to a certain extent, protected theenzyme from inactivation. The results suggest that denaturationof the enzyme protein was involved in inactivation of the enzyme. ¹Part 74 of the phytopathological chemistry of sweet potatowith black rot and injury. ²Present address: Department of Biology, Faculty of Science,Tokyo Metropolitan University, Setagaya-ku, Tokyo. (Received August 5, 1968; )     

Change in potassium distribution in a Phaseolus pulvinus during circadian movement of the leaf

Phaseolus moves its leaves upward and downward with circadianperiod. This movement of the leaf results from the differentialchange in the turgor on opposite sides of the pulvinus. Concentrations of K⁺, Na⁺, Mg⁺⁺, and Ca⁺⁺ in the upper and lowerhalves of the pulvinus and the water content of cells on bothsides of it were analyzed in relation to the deformation ofthe pulvinus. The results showed that (1) the pulvinus was deformedby expansion and contraction of the cells on its opposite sides;(2) among the four cations, the K⁺ concentration was markedlyhigh in both halves of the pulvinus; (3) the osmotic pressureof the upper and lower halves were nearly equal during the rhythmicdeformation of the pulvinus; (4) the expansion and contractionof the cells on the opposite sides of the pulvinus have a positivecorrelation only with a change in the K+ concentration expressedin terms of µmoles per mg protein; (5) the concentrationsof other cations such as Na⁺, Mg⁺⁺, Ca⁺⁺, expressed in termsof µmoles per mg protein, did not change during the circadiandeformation of the pulvinus. Thus, the rhythmic K⁺ movementseems to be the basis for pulvinar turgor movements. With respectto the mechanism of K⁺ movement, three possibilities are discussed. (Received November 7, 1975; )     

Enzymes involved in the last steps of the biosynthesis of mannitol in brown algae

Mannitol-1-phosphate dehydrogenase (EC 1.1.1.17 [EC] ) and mannitol-1-phosphatase(EC number yet unassigned) were detected in the brown algae,Spatoglossum pacificum and Dictyota dichotoma. The enzymes wereextracted from algal fronds and their properties were investigatedusing partially purified preparations. Mannitol-1-phosphatase shows maximum activity at pH 7. The enzymehad a narrow substrate specificity. The Km value for mannitol-1-phosphateis 8.3x10^–4 M (30°C, pH 7.0). The enzyme is activatedby Mg⁺⁺ and Mn⁺⁺and is strongly inhibited by PCMB, Hg⁺⁺and NaF. Mannitol-1-phosphate dehydrogenase showed maximum activitiesat pH values 6.5 and 10.2 in reductive and oxidative reactions,respectively. The dehydrogenase also showed narrow substratespecificity; mannitol-1-phosphate and NAD or fructose-6-phosphateand NADH₂ are utilized, respectively, in oxidative and reductivereactions by the enzyme. Km values for these substrates andthe coenzymes are 2.5x10^–4 M and 7.1x10^–5 M forthe first pair and 2.8x10^–4 M and 1.3x10^–5 M forthe latter pair. This enzyme was strongly inhibited by PCMBand Hg⁺⁺, but was only slightly affected by adenosine phosphates. Possible roles of these enzymes in the biosynthesis of mannitolin brown algae are discussed. ¹ Contributions from the Shimoda Marine Biological Station ofTokyo Kyoiku University, No. 233. This work was supported inpart by a Grant-in-Aid for Co-operative Research from the Ministryof Education, Japan and in part by a grant to one of us (T.Ikawa) from the Matsunaga Science Foundation. ² Present address: Chemical and Physical Laboratory, HoechstJapan Research Laboratory, Minamidai, Kawagoe, Japan. (Received February 22, 1972; )     

Studies on the phosphomannose isomerase of Amorphophallus konjac C. Koch II. Effect of divalent metal ions on the EDTA-treated enzyme

Konjak phosphomannose isomerase was inactivated in a time-dependentprocess by metal binding agents, and the inactivated enzymewas instantaneously reactivated by adding such metal ions asZn⁺⁺, Co⁺⁺, Fe⁺⁺, Mn⁺⁺ and Cu⁺⁺. However, neither Ca⁺⁺ nor Mg⁺⁺were effective for reactivation. Zn⁺⁺, at a low concentration,brought about complete reactivation of the enzyme at pH 6–7. The EDTA-treated enzyme was more susceptible to heat denaturationwhen compared with the native enzyme, but the addition of variousmetal ions caused the recovery of the thermal stability of theEDTA-treated enzyme. The magnitude of the recovery dependedon the metal ion species and the concentrations. The most effectivemetal ion was Co⁺⁺, which caused the recovery of thermal stabilityto a level higher than that of the native enzyme. Phosphomannoseisomerase was inhibited by pchloromercuribenzoate and HgCl₂;the inhibition by p-chloromercuribenzoate being more pronouncedas incubation progressed. In contrast, the EDTA-treated enzymewas more readily inhibited by the mercurial ion than was thenative enzyme. Zn⁺⁺, when added to the EDTA-treated enzyme,markedly restored its resistance to the mercurial-induced inhibition.The metal-substituted enzyme was also inhibited by EDTA in atime-dependent process. ¹ This paper constitutes part 4 of studies on konjak mannanbiosynthesis. (Received March 3, 1975; )     

Electron transport dependent adenosine triphosphatase activity in castor bean endosperm mitochondria

The release of inorganic phosphate from ATP by mitochondriaisolated from endosperms of castor bean (Ricinus communis) wasstimulated by Mg⁺⁺, but not by Ca⁺⁺. EDTA, succinate, NADH₂or oligomycin depressed the reaction. The depression by succinatewas removed by KCN, antimycin A or anoxia. DNP alone did notaffect activity but did stimulate the Pi release in the presenceof succinate under aerobic conditions. Enhanced Pi release inthe presence of succinate and DNP was cancelled by KCN, antimycinA, oligomycin or anoxia. On the basis of these results, themechanism of ATPase action in castor bean endosperm mitochondriais discussed. (Received January 27, 1969; )     

Mg++ -activated and -inhibited ATPases from mung bean hypocotyls

Mg⁺⁺-activated and inhibited ATPases were isolated from dark-grownmung bean hypocotyls. The enzymes hydrolyzed nucleoside tri-,di- and monophosphates and ß-glycerophosphate. Theeffect of Mg⁺⁺ was most marked when ATP and other nucleosidetriphosphates were used as substrates. Mg⁺⁺-activated ATPases: The activity of enzyme-I was localizedin the membranes and was not released by treatment with 0.1%deoxycholate. Enzyme-II was released and separated by CM-cellulosecolumn chromatography. Enzyme-V was separated from the solublefraction of the cell homogenate by DEAE-cellulose column chromatography.The rates of activivation by Mg⁺⁺ of enzyme-II and enzyme-Vwere very small compared to that of enzyme-I. Mg⁺⁺-inhibited ATPases: Enzyme-II and -IV were precipitatedwith 50–80% ammonium sulfate from the soluble fractionof the cell homogenate and were separated by successive columnchromatographies on Sepharose 6B and DEAE-cellulose. The activitiesof enzyme-III and -IV were inhibited by Mg⁺⁺, when ATP, UTPand GTP were used as substrates. Enzyme-III was purified approximately38-fold, and was more remarkably inhibited by Mg⁺⁺ than wasenzyme-IV. ¹Present address: Institute for Plant Virus Research, 959 Aobacho,Chiba 280, Japan. (Received January 7, 1974; )     

Thermal Properties of Membrane Lipids from two Cyanobacteria, Anacystis nidulans and Synechococcus sp.

The composition and positional distribution of fatty acids inmonogalactosyldiacylglycerol, digalactosyldiacylglycerol, phosphatidylglyceroland sulphoquinovosyldiacylglycerol from two cyanobacteria, Anacystisnidulans and Synechococcus sp. grown at 25°C have been determinedand compared with measurements of the phase separation temperaturesof the lipids. Only monogalactosyldiacylglycerol in Anacystisand sulphoquinovosyldiacylglycerol in Synechococcus showed phaseseparation temperatures above 0°C. The phase transitiontemperature of a sample of sulphoquinovosyldiacylglycerol containingover 90% of the dihexadecanoyl molecular species has been determinedto be 43°C for the Na⁺ salt and 38°C for the Mg⁺⁺ salt. ^*Deceased. September 14, 1986. (Received June 25, 1986; Accepted August 25, 1986)     

Further studies on ribulose-l,5-diphosphate carboxylase from Rhodopseudomonas spheroides and Rhodospirillum rubrum

Some enzymic properties of RuDP carboxylase isolated from Rhodopseudomonasspheroides and Rhodospirillum rubrum, grown under autotrophic,semi-autotrophic and heterotrophic conditions were studied.Regardless of the growing condition, the nature of the enzymefrom each respective bacterium remained unchanged. The molecularweight of R. spheroides and R. rubrum RuDP carboxylase was estimatedto be approximately 2.4x10⁵ and 8.3x10⁴, respectively. Bothenzymes require Mg⁺⁺. Their kinetic properties were also examined. ¹Part XII. Structure and Function of Chloroplast Proteins ²Supported in part by research grants from the Ministry of Educationof Japan (No. 8719), USPHS (AM. 10792-03) and the Asahi Press(Tokyo) (Received March 18, 1970; )     

Effects of cations on the sugar receptor-site of the blowfly, Phormia

1) Ca^{+ +} (1 to 10 mM) lowered the binding affinity of sugarreceptor-site for sucrose in the labellar sugar receptor ofthe blowfly, Phormia regina, without changing the maximum-responseamplitude. It also elevated the values of the Hill coefficient(n_H) in some degrees. 2) Other divalent cations such as Mg^{+ +}, Ba^{+ +} or Cd^{+ +} alsoshowed almost the same property as above. The sequence of theeffect is as follows: Ba^{+ +}, Mg^{+ +} x Ca^{+ +} x Cd^{+ +}. Trivalentcation, La^{+ + +} (1 mM), changed the value of n_H from 1 (La^{++ +}-free) to 2. 3) On the contrary, the action of monovalent cations such asK⁺ or Na⁺, of which ionic strength was made the same as thatof the divalents hardly suppressed the response. 4) The results obtained do not support the hypothesis, at leaston the sugar receptor of the fly, that the receptor potentialis attributable to a change of the surface potential (zeta potential)as is proposed for the frog sugar receptor.     

Long chain fatty acid synthesis in developing castor bean seeds I. The operation of the path from acetate to long chain fatty acids in a subcellular particulate system

The path of LFA synthesis from acetate in developing castorbean seeds was associated with subcellular 10,000 g particles.Further fractionation of these particles by a stepwise densitygradient method showed the high possibility that the site ofLFA synthesis is the proplastid. A study on cofactor requirementswhen [1-¹⁴C]acetate predominantly incorporated into LFAs indicatedthat synthesis would be achieved by acetyl-CoA carboxylation,malonyl-ACP condensation. ATP, CoA, HCO₃^– and Mg⁺⁺ orMn⁺⁺ were essential for synthesis from acetate by the 10,000gparticulate system. Results of inhibhitor experiment suggestedthat the supply of ATP to the LFA synthesizing system is broughtabout by mitochondrial oxidative phosphorylation, when acetateis the sole precursor for LFA synthesis in this system. TheNADPH generating system was contained in the paticles, althoughthe addition of NADP⁺ and G-6-P increased synthesis. NADH markedlystimulated LFA synthesis from acetate. The primary role of NADHseems to be as a direct reductant in both steps involving thereduction and oxidative desaturation of fatty acid chains; particularly,in the former step, although NADH partially contributes to thesupply of ATP as a respiratory substrate. It is unlikely thatNADH works as a hydrogen donor to NADP⁺. LFA synthesis by thecastor bean particulate system was not stimulated by light,thus differing from that by leaf chloroplasts. (Received July 23, 1973; )