Distribution of enzymes related to C3 and C4 pathway of photosynthesis between mesophyll and bundle sheath cells of Panicum hians and Panicum milioides

Enzymes of the C₄, C₃ pathway and photorespiration have beenanalyzed for P. hians and P. milioides, which have chlorenchymatousbundle sheath cells in the leaves. On whole leaf extracts thelevels of PEP carboxylase are relatively low compared to C₄species, RuDP carboxylase is typical of C₃ species, and enzymesof photorespiratory metabolism appear somewhat intermediatebetween C₃ and C₄. Substantial levels of PEP carboxylase, RuDPcarboxylase, and photorespiratory enzymes were found in bothmesophyll and bundle sheath cells. Low levels of C₄-acid decarboxylatingenzymes may limit the capacity for C₄ photosynthesis in P. hiansand P. milioides. The results on enzyme activity and distributionbetween mesophyll and bundle sheath cells are consistent withCO₂ fixation via C₃ pathway in these two species. ¹ This research was supported by the College of Agriculturaland Life Sciences, University of Wisconsin, Madison; and bythe University of Wisconsin Research Committee with funds fromthe Wisconsin Alumni Research Foundation; and by the NationalScience Foundation Grant BMS 74-09611. (Received September 16, 1975; )     

Characterization of the Type of Photosynthetic Carbon Dioxide Fixation in Jute (Corchorus olitorius L.)

Activities of photosynthetic and photorespiratory enzymes viz.,ribulose bisphosphate carboxylase, phosphoenol pyruvate carboxylaseand glycolate oxidase from jute (Corchorus olitorius L.; cv.JRO 632) leaves were compared with those from maize (C₄) andsunflower (C₃) leaves. The photosynthetic CO₂ fixation products,the release of ¹⁴CO₂ in light and dark following photosynthesisin ¹⁴CO₂, chlorophyll a: b ratio, gross leaf photosyntheticrate and dry matter production rate were also studied. The resultsshow that jute is a C₃ plant. Key words: Jute, Corchorus olitorius, C₃ photosynthesis     

Ribulose-1,5-Diphosphate Carboxylase Protein during Flag Leaf Senescence

Ribulose-l,5-diphosphate (RuDP) carboxylase protein and activitywere determined in relation to net photosynthetic rate duringthe senescence of intact flag leaves of wheat on the plant.Initially the decrease in RuDP carboxylase activity was greaterthan the decline in net photosynthesis. The major decrease inRuDP carboxylase activity over this period resulted from a decreasein enzyme specific activity from 11 to 2 µmol CO₂ fixedh^–1 mg^–1 protein. Loss of RuDP carboxylase proteindid not occur until late in senescence by which time chlorophyllconcentration had decreased by more than 50%. Treatment of flagleaves at weekly intervals with either 1000 parts 10^–62-chloro-ethyltrimethylammonium chloride or 100 parts 10^–6gibberellic acid with 1 part 10^–6 kinetin did not significantlyaffect net photosynthetic rate, RuDP carboxylase protein oractivity during senescence.     

CO2-Fixation, Glycolate Formation, and Enzyme Activities of Photorespiration and Photosynthesis during Greening and Senescence of Sunflower Cotyledons

Time-courses of ¹⁴CO₂-fixation and of enzyme activities involvedin photorespiration and photosynthesis were determined duringthe life span of cotyledons from sunflower seedlings (Helianthusannuus L.). Glycolate formation in vivo was estimated from theresults of combined labelling and inhibitor experiments. NADPH-glyceraldehyde-3-phosphatedehydrogenase, NADPH-glyoxylate reductase and chlorophyll werewell correlated with the time-course of ¹⁴CO₂-fixation (photosynthesis).There was, however, a considerable discrepancy between the developmentalsequence of photosynthesis and that of both ribulose-l,5-bisphosphatecarboxylase and glycolate oxidase. Furthermore, time-coursesof glycolate oxidase activity in vitro and of glycolate formationin vivo differed significantly. Therefore, the use of glycolateoxidase as a marker for the activity of photorespiration ingreening sunflower cotyledons may be questionable. Results from¹⁴CO₂-labelling experiments with cotyledons treated with theglycolate oxidase inhibitor 2-hydroxy butynoic acid suggestthat glycolate formation relative to CO₂-fixation is reducedin senescent cotyledons. Key words: Development, glycolate oxidase, photorespiration, ribulose-l,5-bisphosphate carboxylase, oxygenase     

Glycine as a substrate for photorespiration

Substrates for photorespiration were examined by feeding ¹⁴Clabeled compounds to tobacco and corn leaf segments and by measuring¹⁴CO₂ evolution in light and darkness. CO₂ release in the darkwas rapid, but in light CO₂ release was slow due to refixationby photosynthesis. Carboxyl labeled glycine was more rapidlydecarboxylated than were glyoxylate, glycolate or serine. Hydroxypyridinemethanesulfonate, an inhibitor of glycolate oxidase, blocked CO₂ releasefrom glycolate but not from glycine. Isonicotynyl hydrazideblocked CO₂ release from both glycine and glycolate. DCMU blockedphotosynthetic refixation of the released CO₂, consequentlythe rates of CO₂ release in light and dark were about equal.It was concluded that CO₂ release during photo-respiration camefrom the conversion of 2 molecules of glycine to one serineand one CO₂. ¹⁴CO₂ release from glycine-l-¹⁴C in the dark or with DCMU inlight can be used as an assay for photorespiration ability. CO₂ release from glycine and glycolate by corn leaf segmentsin the dark proceeded at the rate of that in normal tobaccoleaf. This result, together with other work on O₂ exchange andenzymatic analysis, indicates that corn and other plants docarry on photorespiration, but it is not manifested by CO₂ releasein light. A yellow tobacco mutant, Consolation 402, had high rates ofphotorespiration by the ¹⁴CO₂ assay, nearly half (or more) asmany peroxisomes as chloroplasts, and high rates of CO₂ releasefrom glycine-l-¹⁴C or glycolate-l-¹⁴C. A common tobacco, BrightYellow, had lower rates of photorespiration, fewer visible peroxisomes,and slower decarboxylation of glycine and glycolate. The amount of ¹⁴CO₂ release from glycine-l-¹⁴C or glycolate-l-¹⁴Cincreased only slightly when the temperature was raised from25 to 35°C. ¹Parts of this work were abstracted at the Annual Meeting (April,1969) of Japanese Society of Plant Physiologists, Kanazawa ²Department of Biochemistry, Michigan State University, EastLansing, Michigan, U.S.A. (Received September 3, 1969; )     

Photosynthetic efficiency of Panicum hians and Panicum milioides in relation to C3 and C4 plants

Panicum hians and Panicum milioides were found to have characteristicsintermediate to those of C₃ and C₄ species with respect to CO₂compensation point, percentage inhibition of photosynthesisby O₂ at various O₂/CO₂ solubility ratios, and water use efficiency.C₄ species have a higher carboxylation efficiency than eitherthe intermediate or C₃ species. During photosynthesis, evenunder 2.5% O₂, C₄ species have a higher affinity for intercellularCO₂ (Km 1.6 µM) apparently due to the initial carboxylationthrough PEP carboxylase. Under low O₂ the intermediate and C₃species had a similar affinity for intercellular CO₂ duringphotosynthesis (Km 5–7 µM) consistent with carboxylationof atmospheric CO₂ through RuDP carboxylase. There were considerablevariation in photosynthesis/unit leaf area at saturating CO₂levels in the species examined which in part is due to differencesin RuDP carboxylase /unit leaf area. The highest rates of photosynthesis/unitleaf area under CO₂-saturating conditions were with the C₃ specieswhich had a correspondingly high level of RuDP carboxylase/unitleaf area. Possibilities for the greater efficiency of P. hiansand P. milioides in comparison to C₃ species in utilizing lowlevels of CO₂ in the presence of atmospheric O₂ are discussed. ¹ This research was supported by the College of Agriculturaland Life Sciences, University of Wisconsin, Madison; and theUniversity of Wisconsin Research Committee with funds from theWisconsin Alumni Research Foundation. (Received June 25, 1977; )     

A CO2 Concentrating System in Leaves of Higher C3-Plants Predicted by a Model Based on RuBP Carboxylase/Oxygenase Kinetics and 14CO2/12CO2 Exchange

Mächler, F., Lehnherr, B., Schnyder, H. and Nösberger,J. 1985. A CO₂ concentrating system in leaves of higher C₃-plantspredicted by a model based on RuBP carboxylase/oxygenase kineticsand ¹⁴CO₂/¹²CO₂ exchange.–J. exp. Bot. 36: 1542–1550. A model is presented which compares the ratio of the two activitiesof the enzyme nbulose bisphosphate carboxylase/oxygenase asdetermined in vitro with the ratio of photosynthesis to photorespirationin leaves as determined from differential ¹⁴CO₂/¹²CO₂ uptakeor from CO₂ compensation concentration. Discrepancies betweenmeasurements made in vitro and in vivo are attributed to theeffect of a CO₂ concentrating system in the leaf cells. Interferencefrom dark respiration is discussed. A CO₂ concentrating systemis postulated which is efficient mainly at low temperature andlow CO₂ concentration. Key words: —Photosynthesis, photorespiration, ribulose bisphosphate carboxylase/oxygenase     

The Mechanism of Photosynthesis in the Tea Plant (Camellia sinensis L.)

Leaves of the tea plant photosynthesizing in ¹⁴CO₂ incorporatedmuch radioactivity into intermediates of the glycolate pathwayand little into C₄ acids. Increased O₂ in the atmosphere decreasedphotosynthesis, stimulated photorespiration, and increased theCO₂ compensation point. In air the rate of photorespirationwas 19% of net photosynthesis. These observations indicate aC₃ rather than a C₄ mechanism of photosynthesis.     

Changes in the Rate of Photosynthesis during Grain Filling and the Enzymatic Activities Associated with the Photosynthetic Carbon Metabolism in Rice Panicles

Photosynthesis is known to occur in rice panicles, but littlehas been reported about the photosynthetic or biochemical characteristicsof such panicles. The estimated gross amount of photo-syntheticallyassimilated CO₂ in a panicle is 30% of that in a flag leaf.This result and the good light-intercepting characteristicsof the panicle in the canopy suggest that photosynthesis inthe panicle may contribute significantly to grain filling. Therice panicle is composed of spikelets and of rachis-branchesincluding rachis which have estimated gross rates of photosynthesisduring the 30-day period after anthesis of 130 to 180 and 50to 100 µmol CO₂.(mg Chl)^–1.h^–1, respectively.The corresponding rate for the flag leaf is 180 to 230 µmolCO₂.(mg Chl)^–.h^–. On the basis of Chl, spikeletshave a high photosynthetic capability which is similar to thatof the flag leaf. The activities of ribulose-l,5-bisphosphate carboxylase (RuBPCase),phosphoenolpyruvate carboxylase (PEPCase), and pyruvate.P_i dikinase(PPDK) in spikelets were 129, 220, and 87 µmol.(mg Chl)^–.h^–,respectively. The activities of PEPCase and PPDK in spikeletswere considerably higher than those in the flag leaf or rachis-branches.Oxygen-insensitive photosynthesis was found only in spikelets.The K_m of NaHCO₃ for photosynthesis by slices of spikelets inan aqueous solution (0.6 mM) was considerably lower than thatfor slices of flag leaf (4.2 mM). All these results indicatethat spikelets have different photosynthetic characteristicsfrom those of the flag leaf and rachis-branches. The possibilityof C₃–C₄ intermediate photosynthesis or C₄-like photosynthesisin spikelets is discussed. ⁴Present address: Department of Biochemistry, Faculty of Science,Saitama University, Urawa, 338 Japan (Received February 14, 1990; Accepted June 12, 1990)     

Metabolism of epidermal tissues, mesophyll cells, and bundle sheath strands resolved from mature nutsedge leaves

Mesophyll cells and bundle sheath strands were isolated from Cyperus rotundus L. leaf sections infiltrated with a mixture of cellulase and pectinase followed by a gentle mortar and pestle grind. The leaf suspension was filtered through a filter assembly and mesophyll cells and bundle sheath strands were collected on 20-μm and 80-μm nylon nets, respectively. For the isolation of leaf epidermal strips longer leaf cross sections were incubated with the enzymes and gently ground as above. Loosely attached epidermal strips were peeled off with forceps. The upper epidermis, which lacks stomata, could be clearly distinguished from the lower epidermis which contains stomata. Microscopic evidence for identification and assessment of purity is provided for each isolated tissue.Enzymes related to the C₄-dicarboxylic acid cycle such as phosphoenolpyruvate carboxylase, malate dehydrogenase (NADP⁺), pyruvate, P_i dikinase were found to be localized, ≥98%, in mesophyll cells. Enzymes related to operating the reductive pentose phosphate cycle such as RuDP carboxylase, phosphoribulose kinase, and malic enzyme are distributed, ≥99%, in bundle sheath strands. Other photosynthetic enzymes such as aspartate aminotransferase, pyrophosphatase, adenylate kinase, and glyceraldehyde 3-P dehydrogenase (NADP⁺) are quite active in both mesophyll and bundle sheath tissues.Enzymes involved in photorespiration such as RuDP oxygenase, catalase, glycolate oxidase, hydroxypyruvate reductase (NAD⁺), and phosphoglycolate phosphatase are preferentially localized, ≥84%, in bundle sheath strands.Nitrate and nitrite reductase can be found only in mesophyll cells, while glutamate dehydrogenase is present, ≥96%, in bundle sheath strands.Starch- and sucrose-synthesizing enzymes are about equally distributed between the mesophyll and bundle sheath tissues, except that the less active phosphorylase was found mainly in bundle sheath strands. Fructose-1,6-diP aldolase, which is a key enzyme in photosynthesis and glycolysis leading to sucrose and starch synthesis, is localized, ≥90%, in bundle sheath strands. The glycolytic enzymes, phosphoglyceromutase and enolase, have the highest activity in mesophyll cells, while the mitochondrial enzyme, cytochrome c oxidase, is more active in bundle sheath strands.The distribution of total nutsedge leaf chlorophyll, protein, and PEP carboxylase activity, using the resolved leaf components, is presented. ¹⁴CO₂ Fixation experiments with the intact nutsedge leaves and isolated mesophyll and bundle sheath tissues show that complete C₄ photosynthesis is compartmentalized into mesophyll CO₂ fixation via PEP carboxylase and bundle sheath CO₂ fixation via RuDP carboxylase. These results were used to support the proposed pathway of carbon assimilation in C₄-dicarboxylic acid photosynthesis and to discuss the individual metabolic characteristics of intact mesophyll cells, bundle sheath cells, and epidermal tissues.     

Influence of Temperature on the Ratio of Ribulose Bisphosphate Carboxylase to Oxygenase Activities and on the Ratio of Photosynthesis to Photorespiration of Leaves

Lehnherr, B., Mächler, F. and Nösberger, J. 1985.Influence of temperature on the ratio of ribulose bisphosphatecarboxylase to oxygenase activities and on the ratio of photosynthesisto photorespiration of leaves.—J. exp. Bot. 36: 1117–1125. Rates of net and gross photosynthesis of intact white cloverleaves were measured by infrared gas analysis and by short termuptake of ¹⁴CO₂ respectively. Ribulose bisphosphate carboxylaseoxygenase (RuBPCO) was purified from young leaves and kineticproperties investigated in combined and separate assays. Theratio of carboxylase to oxygenase activities was compared withthe ratio of photosynthesis to photorespiration at various temperaturesand CO₂ concentrations. The ratio of photosynthesis to photorespiration at 30 Pa p(CO₂)was consistent with the ratio of carboxylase activity to oxygenaseactivity when each was measured above 20 °C. However, theratio of photosynthesis to photorespiration increased with decreasingtemperature, whereas the ratio of carboxylase to oxygenase activitywas independent of temperature. This resulted in a disagreementbetween the measurements on the purified enzyme and intact leafat low temperature. No disagreement between enzyme and leafat low temperature occurred, when the ratio of photosynthesisto photorespiration was determined at increased CO₂ concentrations. The results suggest an effect of low temperature and low CO₂concentration on the ratio of photosynthesis to photorespirationindependent of the enzyme. Key words: Ribulose bisphosphate carboxylase oxygenase, photorespiration, temperature     

Photosynthetic carbon metabolism in wild, primitive and cultivated forms of wheat at three levels of ploidy: Role of the glycolate pathway

¹⁴CO₂ assimilation was studied with diploid, tetraploid, hexaploidspecies of the genera Triticum and their wild relatives Aegilops.Attached mature leaves of 3–4 weekold plants were allowedto undergo photosynthesis under air at ambient temperature.The pattern of distribution of ¹⁴C was notably similar in Triticumand Aegilops species whatever the level of ploidy. Sucrose wasthe sink for photosynthetic carbon. ¹⁴C for sucrose synthesis was supplied either through the glycolatepathway by glycolate, the product of the photorespiration orby the Calvin cycle intermediates exported into the cytoplasm.Depending on the species, the glycolate pathway provided 40to 75%of the sucrose ¹⁴C. The higher labeling of sucrose was associated with the greaterparticipation of the glycolate pathway in the wild diploid (DD)A. squarrosa and in the cultivated hexaploid (AABBDD) T. aestivum.The results suggest that the expression of the male D genomeis dominant over the female AB genome in T. aestivum. In T. aestivum under ambient conditions lowering (low temperature)or hindering (1% O₂ ) photorespiration, sucrose labeling decreased,but serine and glycine labeling was favoured. We propose thatin wheat leaves, the role of photorespiration is to drain artof the carbon exported from the chloroplast as glycolate, towardssucrose synthesis. (Received March 16, 1979; )     

Relationship between leaf development,carboxylase enzyme activities and photorespiration in the C4-plant Portulaca oleracea L.

Summary Ribulose diphosphate (RuDP) and (PEP) phosphoenolpyruvate carboxylase enzyme activities were studied in young, mature, and senescent Portulaca oleracea leaves. While the absolute amount of both the C₃ (RuDP) and C₄ (PEP) carboxylase is less in senescent leaves than in mature leaves, RuDP carboxylase activity is reduced to a lesser degree. In senescent leaves, PEP carboxylase activity equals 10% of that in mature tissue, but RuDP carboxylase is 27% of that in mature leaves. The same ontogenetic series was also used to determine photorespiration rates and responses to several gas treatments. Young and mature leaves were unaffected by changes in the light regime or oxygen concentrations, and exhibited typical C₄-plant light/dark ¹⁴CO₂ evolution ratios. Senescent leaves, on the other hand, have photorespiration ratios similar to C₃-plants. In addition, senescent leaves were affected by minus CO₂, 100% O₂ and N₂ in a manner expected of C₃-plants, but not C₄-plants. These results are discussed in terms of a relative increase in activity of the C₃ cycle in later developmental stages in this plant.Abbreviation RuDP ribulose diphosphate - PEP phosphoenolpyruvate - PGA phosphoglyceric acid     

Comparison of carbon dioxide fixation and the fine structure in various assimilatory tissues of Amaranthus retroflexus L

The pattern for primary products of CO₂-fixation and the chloroplaststructure of Amaranthus retrqflexus L., a species which incorporatescarbon dioxide into C₄ dicarboxylic acids as the primary productof photosynthesis, were compared in various chlorophyll containingtissues,i.e., foliage leaves, stems, cotyledons and pale-greencallus induced from stem pith. Despite some morphological differencesin these assimilatory tissues, malate and aspartate were identifiedas the major compounds labelled during a 10 sec fixation of¹⁴CO₂ in all tissues. Whereas, aspartate was the major componentin C₄-dicarboxylic acids formed in foliage leaves, malate predominatedas the primary product in stems, cotyledons and the pale-greencallus. The percentage of ¹⁴C-radioactivity incorporated intoPGA and sugar-P esters increased and ¹⁴C-sucrose was detectedin the prolonged fixation of ¹⁴CO₂ in the light, not only infoliage leaves, but also in stems and cotyledons. ¹ This work was supported by a Grant for Scientific ResearchNo. 58813, from the Ministry of Education, Japan. ² Present address: Institute of Applied Microbiology, Universityof Tokyo, Tokyo, Japan. ³ Present address: Department of Biochemistry, University ofGeorgia, Athens 30601. Georgia, U. S. A. (Received July 10, 1971; )     

Photoinhibition under Atmospheric O2, the Activation State of RuBP Carboxylase and the Content of Photosynthetic Intermediates in Soybean and Wheat

Ward, D. A. and Drake, B. G. 1987. Photoinhibition under atmosphericO₂, the activation state of RuBP carboxylase and the contentof photosynthetic intermediates in soybean and wheat.—J.exp. Bot. 38: 1937–1948. Associations between photosynthesis, the activation state ofRuBP carboxylase and the contents of photosynthetic intermediateswere compared in soybean and wheat leaves before and after exposureto photoinhibitory treatments in the presence of atmosphericO₂. Exposing attached leaves to a supra-saturating irradiance(3 800 µmol quanta m^{– 2} s^–1) for 2 h in CO_2-freeair decreased carboxylation efficiency and the light-saturatedphotosynthetic rate in air by approximately 50%. Exposure tothe photoinhibitory treatment for periods in excess of 2 h didnot cause a further decrease of photosynthesis in soybean. Althoughphotosynthesis was reduced, the initial and total (fully-activated)activities of ribulose 1,5-bisphosphate carboxylase (RuBPCase)in leaf extracts were unaltered in each species by the photoinhibitorytreatment. This was true for leaves sampled under both air andat a rate-limiting intercellular CO₂ partial pressure (C_i) of75 µPa Pa^–1. The contents of ribulose l,5-bisphosphate(RuBP) and 3-phosphoglyceric acid (3-PGA) were reduced by thephotoinhibitory treatment in soybean leaves sampled in air andat a rate-limiting C_i, although the RuBP/3-PGA ratio was unaffected.The relative reduction of RuBP content in soybean leaves atrate-limiting C₁ was similar to the corresponding reductionof carboxylation efficiency. For wheat,the relative reductionof RuBP content at rate-limiting C_i (–19%) caused by thephotoinhibitory treatment was considerably less than the correspondingdecrease of carboxylation efficiency (–49%).The RuBP/3-PGAratio of wheat was also increased significantly by the photoinhibitorytreatment The significance of these observations to the regulationof CO₂-limited photosynthesis in leaves experiencing photoinhibitionunder atmospheric oxygen is discussed. Consideration is alsogiven to the previous contention that contemporary measurementsof initial activity in crude extracts may provide a spuriousindication of the amount of the enzyme-CO₂-Mg_{2 +} form of RuBPcarboxylase present in the leaf. Key words: Carboxylation efficiency, RuBP carboxylase, photoinhibition, RuBP, 3-PGA     

Compartmental modelling of photorespiration and carbon metabolism of water stressed leaves

Abstract Carbon fluxes in photosynthesis and photorespiration of water stressed leaves have been analysed in a steady state model based on the ribulose diphosphate carboxylase (RuDP carboxylase) and RuDP oxygenase enzyme activities and the CO₂ and O₂ concentrations in the leaf. Agreement between predicted and observed photorespiration (Lawlor & Fock, 1975) and C flux in the glycollate pathway is good over much of the range of water stress, but not at severe stress. An alternative source of respiratory CO₂ is suggested to explain the discrepancy. The model suggests that resistance to CO₂ fixation is mainly in the carboxylation reactions, not in CO₂ transport. Using the steady state model, the kinetics of ¹⁴C incorporation into photosynthetic and photorespiratory intermediates are simulated. The predicted rate of ¹⁴C incorporation is faster than observed and delay terms in the model are used to simulate the slow rates of mixing and metabolic reactions. Inactive pools of glycine and serine are suggested to explain the observed specific activities of glycine and serine. Three models of carbon flux between the glycollate pathway, the photosynthetic carbon reduction cycle and sucrose synthesis are considered. The most satisfactory simulation is for glycollate pathway carbon feeding into the PCR cycle pool of 3-phosphoglyceric acid which provides the carbon for sucrose synthesis. Simulation of the specific activity of CO₂ released in photorespiration suggests that a source of unlabelled carbon may contribute to photorespiration.     

Glycolate synthesis in a C3 C4 and intermediate photosynthetic plant type

Excised leaves of a C₃-photosynthetic type, Hordeum vulgare,a C₄-type, Panicum miliaceum, and an intermediate-type, Panicummilioides, were allowed to take up through their cut ends a1 mM solution of butyl hydroxybutynoate (BHB), an irreversibleinactivator of glycolate oxidase. After 30 to 60 min in BHB,extractable glycolate oxidase activity could not be detectedin the distal quarter of the leaf blades. Following this pretreatment,recovery of ¹⁴C-glycolate from ¹⁴CO₂ incorporated in a 10 minperiod was nearly maximal for each of the three plant types.Labeled glycolate was 51% of the total ¹⁴CO₂ incorporated forthe C₃-species, 36% for the intermediate-species, and 27% forthe C₄-species Increased labeling of glycolate was compensatedfor primarily by decreased labeling of the neutral and basicfractions for the C₃ and intermediate-type species. In the C₄-type,label decreased primarily in the neutral and insoluble fractions,but increased in the basic fraction. A lower rate of glycolatesynthesis is indicative of a lower rate of photorespirationand consistent with a lower O₂/CO₂ ratio present in the bundle-sheathcells of C₄-plants. We conclude that both decreased glycolatesynthesis and the refixation of photorespiratory-released CO₂are important in maintaining a lower rate of photorespirationin C₄-plants compared to C₃ plants. Intermediate glycolate synthesisin Panicum milioldes is consistent with its intermediate levelof O₂ inhibition of photosynthesis and intermediate rate ofphotorespiration. (Received May 6, 1978; )     

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; )     

Photosynthesis, Respiration, and Carbon Assimilation in Water-Stressed Maize at Two Oxygen Concentrations

Photosynthesis decreased with decreasing leaf water potentialas a consequence of stomatal closure and possibly non-stimataleffects of severe stress. Assimilation ceased at c. 16x 10⁵Pa. Photo-respiration, in 21% O₂, was small in relation to assimilationin unstressed leaves and decreased as leaf water potential fellbut it was much larger in proportion to photosynthesis at severestress. Decreasing the O₂ content to 1.5% increased photosynthesisslightly and decreased photo-respiration but did not changethe stress at which assimilation stoped. Dark respiration wasinsensitive to both O₂ and stress. Less ¹⁴C accumulated in stressedleaves but in 21% O₂ a greater proportion of it was in aminoacids, particularly glycine and serine. 1.5% O₂ decreased the¹⁴C in glycine to 10% and in serine to 50% of their levels in21% O₂. In both O₂ concentrations the proportion of ¹⁴C in serineincreased only at the most severe stress. Gas exchange measurementsand changes in the ¹⁴C flux to glycine are interpreted as theresult of glycolate pathway metabolism increasing as a proportionof assimilation in stressed leaves in high O₂. The small absoluterate of photorespiration in high O₂ and at low leaf water potentialmay be due to slow rates of glycine decarbodylation as wellas efficient fixation of any CO₂ produced. Serine is synthesizedby an O₂-sensitive pathway and an O₂-insensitive pathway, whichis most active at severe stress. Synthesis of alanine competeswith that of glycine and serine for a common precursor suppliedby the photo-synthetic carbon reduction cycle. The relativespecific radioactivities of aspartate and alanine suggest thatthey are derived from a common precursor pool, probably pyruvatefrom 3-PGA. The amounts of 3-PGA, aspartate, malate, alanine,and sucrose decreased with increasing water stress as a consequenceof slower assimilation and pool filling. Other amino acids,glycine, serine, glutamate, and proline, accumulated at lowwater potential possibly due to increased synthesis and slowerrates of consumption. Changes in pool sizes, carbon fludes,and specific activities of metabolites are related to the mechanismof C₄ photosynthesis and current concepts of glycolate pathwaymetabolism.     

Effects of growth temperature on photosynthetic carbon metabolism in green plants III. Differences in structure, photosynthetic activities and activities of ribulose diphosphate carboxylase and glycolate oxidase in leaves of wheat grown under varied temperatures

The rate of ferricyanide photoreduction in broken chloroplastsisolated from leaves of wheat acclimatized to a low temperature(mean temperature, 5–7?C) was similar to that in chloroplastsfrom wheat acclimatized to a high temperature (20–25?C). There was no practical difference in glycolate oxidase activityin leaf extracts of wheat plants grown at low and high temperatures.In contrast, the ribulose diphosphate carboxylase activity ofchloroplasts from low temperature sample was less than halfthat for the high temperature sample. Chloroplasts having a high rate of photosynthetic CO₂-fixationwere obtained from wheat acclimatized to a low temperature,whereas the CO₂-fixation activity in chloroplasts isolated fromhigh temperature-acclimatized wheat was very low. Electron microscopy revealed that chloroplasts in high temperature-acclimatizedwheat were ellipsoidal, electron dense and contained starchgranules. Those in low temperature-acclimatized leaves wereround and did not contain starch granule. ²Present address: Department of Botany, Faculty of Science,University of Tokyo, Tokyo, Japan (Received August 7, 1973; )