ONTOGENY AND DIFFERENTIATION OF SCLEREIDS IN RAUWOLFIA

An interesting anatomic feature of Rauwolfia is the occurrence of a remarkable type of sclereid in the stem and root. The initials of the sclereids in the stem arise in the ground tissue element of the pith in a region between 50 and 70μ below the surface of the shoot apex. This region of the shoot remains surrounded by a whorl of either 3 or 4 leaves. Sclereids initiate in succession in association with each whorl of leaves. Thus, the sclereids are restricted to the nodes. The sclereids in the stem arise as a primary element of the shoot from the ground tissue of the pith. In the root, they differentiate from the cells of the phelloderm and are secondary in origin. Morphologically, the sclereids in these 2 organs are basically the same, except that the sclereids in the stem are larger in size than those in the root. A solitary cell, or 2 to several cells in a longitudinal cell file (originated from a single mother cell), may differentiate into sclereid initials. The growth of the sclereids through relatively compact ground tissue of the pith is possibly accomplished by a combination of gliding growth and apical intrusive process. The sclereid initials grow rapidly and force their way between the parenchymatous cells. As a result, the neighboring cells lose their original surface contacts. Sclereids increase in size rapidly, and, therefore, very enlarged sclereids with thin primary walls may be observed in the second node. They mature progressively in basipetal direction in the subjacent nodes. In the fifth or sixth node, mature sclereids with massive secondary walls are most common. The secondary walls of sclereids contain much lignin as determined by the phloroglucinol-HCl test. The walls of sclereids at maturity show a variable number of lamellae ranging from 10 to 15 in the lateral walls. A remarkable feature of the sclereids is their canal-like pits in the secondary walls. Two adjacent pits may coalesce uniquely to form a Y-like configuration directed centrifugally from the lumen of the sclereids. The sclereids are ventrically symmetrical, joined end-to-end by their transverse walls like 2 superimposed young fibers.     

Physiological Diversity in Metabolism in Marine and Terrestrial Crustacea

Two aspects of metabolic adaptation to increased terrestrialismare considered: (1) respiratory adaptations as reflected bycomparative cytochrome c oxidase activity in tissues of crabsfrom aquatic and terrestrial habitats, and (2) thermal acclimationpatterns in cytochrome c oxidase activity in tissues from thesecrabs. Enzymatic assays were done spectrophotometrically ongill, muscle, and mid-gut gland tissues from two aquatic species,Libinia emarginata and Callinectes sapidus, and the terrestrialOcypode quadrata. Cytochrome c oxidase was chosen for this studysince it is generally believed that the more aerobic the cellsor tissues become, the more fully developed the cytochrome systemwill be. This enzyme is also thought to have a role in thermalacclimation. In gill tissue the activity of cytochrome c oxidase is enhancedwith the advent of aerial respiration. Enzymatic activity ofgill tissue from Ocypode quadrata was significantly greaterthan it was in tissue from the aquatic species. No correlationwas observed with increased terrestrialism and enzymatic activityof muscle or mid-gut gland tissue. The thermal acclimation patternsof tissues of these three species of crabs indicate a clear-cuttendency for less enzymatic adaptation to temperature at thetissue level as these crabs evolve toward a land habitat.     

Purification of Sulphite-Cytochrome c Reductase of Thiobacillus novellus and Reconstitution of Its Sulphite Oxidase System with the Purified Constituents

Sulphite-cytochrome c reductase (sulphite: ferricytochrome coxidoreductase, EC 1.8.2.1 [EC] ) derived from Thiobacillus novelluswas purified by chromatography on a DEAE-cellulose column andby gel filtration with a Sephadex G-100 column. Although thereductase thus purified moved as a single band both in gel filtrationand in isoelectric focusing it was always split into two bandsby polyacrylamide gel electrophoresis; the one had the enzymaticactivity and showed absorption spectrum of cytochrome, whilethe other had no activity and was colourless, in contrast withthe results reported by Charles and Suzuki [(1966) Biochim.Biophys. Acta 128: 522]. The enzymatic properties of the purifiedreductase were almost the same as those of the enzyme obtainedby Charles and Suzuki. Cytochrome c-551 free of the reductase activity was obtained.Its molecular weight was determined to be 23,000 by polyacrylamidegel electrophoresis in the presence of sodium dodecyl sulphate.The cytochrome seemed to exist in the organism as a complexwith the reductase or a subunit of the enzyme. In the stateof the complex with the enzyme, the cytochrome was reduced veryquickly on addition of sulphite, while the cytochrome free ofthe reductase activity was hardly reduced by the enzyme withsulphite. A sulphite oxidase system was reconstituted with the reductase,cytochrome c-550 and cytochrome oxidase highly purified fromthe bacterium. ¹ Present address: Water Research Institute, Nagoya University,Nagoya 464, Japan ² Present address: Institute for Biological Science, SumitomoChemical Co., Ltd., Takarazuka, Hyogo 665, Japan (Received January 23, 1981; Accepted March 9, 1981)     

SCLEREID DISTRIBUTION IN THE LEAVES OF PSEUDOTSUGA UNDER NATURAL AND EXPERIMENTAL CONDITIONS

Al -talib , Khalil H., and John G. Torrey . (U. California, Berkeley.) Sclereid distribution in the leaves of Pseudotsuga under natural and experimental conditions. Amer. Jour. Bot. 48(1): 71–79. Illus. 1961.—A study of the distribution of sclereids in cleared leaves taken from 1-, 2-, and 4-year-old shoots of an adult tree of Pseudotsuga menziesii (Mirb.) Franco showed a repeated pattern of sclereid distribution along the shoot axis with many sclereids in the basal leaves grading into few or no sclereids in the terminal leaves of each year's growth. Attempts were made to influence sclereid distribution by bud defoliation of attached branches with and without auxin treatment and by testing the effects of growth-regulating substances on sclereid formation in leaves of excised buds of Pseudotsuga cultured in vitro. Whereas removal of the basal ¾ of the leaves at the time of bud unfolding had no effect on bud, leaf or sclereid development, removal of the leaves of the upper half or complete defoliation led to premature expansion of next year's terminal bud with leaves developing in part from presumptive bud-scale primordia. Indoleacetic acid at 0.5% in lanolin paste applied to the defoliated region prevented this premature bud expansion. Defoliation of the basal half did not affect sclereid formation in the terminal leaves. Sclereid development in leaves of prematurely expanded buds on defoliated branches was normal except in the few cases where bud expansion occurred in the presence of low-auxin concentrations. Then, sclereid development was inhibited. Sclereid formation in leaves of excised buds grown in nutrient culture was generally much less frequent than in intact branches, and auxin treatment still further reduced the frequency of sclereids. It was concluded that sclereid initiation and differentiation in the intact plant may well be under the control of hormonal factors in the plant, one of which may be auxin.     

Effects of Developing Leaves on Stelar Pattern Development in the Shoot Apex of Matteuccia struthiopteris

A developmental study of the normal shoot apex of Matteucciastruthiopteris suggested that patterned stelar differentiationis initiated immediately beneath the single layer of promeristemand occurs prior to the initiation of the youngest leaf primordium.A developmental study in which all leaf primordia were suppressed,with or without lateral isolation of the terminal meristem byvertical incisions, has confirmed this interpretation of stelardifferentiation. Experimentally-induced changes in the tissueimmediately below the promeristem were reflected in the resultingmature structure of the stele. Failure of leaf gap initialsto differentiate, if all leaf primordia were suppressed at theincipient stage, resulted in a mature stele without leaf gaps.Similarly the disappearance of pith mother cells after severalweeks of leaf removal was associated with the formation of astele without pith. Leaf influence was further assessed by allowingone primordium to develop while all others were suppressed.The developing leaf had a small promoting effect on caulinevascular tissue differentiation but its major impact on theexpansion of the parenchymatous tissues of the stele. Characteristicprotoxylem and protophloem failed to differentiate when allleaves were suppressed and, when leaf was allowed to develop,formed only in relation to the leaf.Copyright 1995, 1999 AcademicPress Leaf influence, vascular pattern formation, experimental surgery, shoot apex development, protoxylem, protophloem, Matteuccia struthiopteris     

The Development of the Fruit Walls in Carya

Both the pistillate flowers and the developing fruits of CaryaovataandC. cordiformis are covered with secretory and protectivehairs. There are two kinds of the secretory hairs. In the morefrequent one the volatile oils are accumulated under the cuticlecovering the cells of the head. In the other the secretory substancesare accumulated inside the cells. The flowers and the developingfruits of the two investigated species have quite large emergenceson which the stomata most often occur. During the developmentof the fruit lenticels are formed in the green hull. Tanninsare accumulated not only in the green hull, but also in thecarpellary layer and in the packing tissue. There are some differences between C. ovata and C. cordiformisin the course of sclerification of the green hull. In C. cordiformisthe hull is less sclerified than in C. ovata, and the sclerificationof the carpellary layer occurs later. At a certain period ratherlarge gaps are present in the lignifying shell (in the regionsof the sutures) where the cells have thin cellulose walls. Thesegaps are partly filled with sclereids. The gaps become reducedto narrow chinks, in which the cell walls remain unlignified.The hard shell is not formed in all its width at the same time.During the process of formation of the hard fruit shell theremay arise single sclereids surrounded by thin-walled cells.Within a short time these undergo a similar modification andthe tissue becomes uniform as sclerenchyma. The sclereids ofthe hard shell have, in general, strongly folded cell walls.Owing to this they overlap each other.     

Studies on the change of the respiratory system in roots in relation to the growth of plants II. Activity of iron-containing oxidases in roots of cereal crops with the aging of plants

Distribution of iron-containing oxidases in aging nodal rootsof rice and wheat was studied. Activities of cytochrome c oxidase(1.9.3.1 [EC] , cytochrome c : O₂ oxidoreductase), catalase (1.11.1.6 [EC] ,H₂O₂: H₂O₂ oxidoreductase) and peroxidase (1.11.1.7 [EC] , donor:H₂O₂ oxidoreductase) in wheat roots were comparatively higherthan were those in rice roots at corresponding stages. Cytochromec oxidase in roots remained active throughout the lives of therice and wheat crops. In rice roots, catalase seemed to playa distinct role around the panicle formation stage. Decay ofcatalase activity took place earlier than did that of peroxidaseand cytochrome c oxidase activities. In wheat roots similarenzyme activity changes were not observed. Data may suggestthat the high activity of iron containing oxidases at the panicleformation stage (I) may be chiefly due to catalase activityin rice roots. ¹Paper presented at the 14th Annual Meeting of the Society ofthe Science of Soil and Manure, Japan (1968). (Received November 21, 1968; )     

Vascular Differentiation in the Shoot Apex of Matteuccia struthiopteris

Initial vascular differentiation is generally considered tooccur in procambium. In ferns, however, a provascular tissueimmediately subjacent to the promeristem has been suggestedas an initial stage within which the procambium is subsequentlyformed. In contrast to this interpretation, a zonation conceptapplied in ferns recognizes a promeristem consisting of severallayers of cells in which no differentiation takes place. Thisstudy demonstrates that the shoot apex of Matteuccia struthiopterishas one cell layer of promeristem. Immediately subjacent tothe promeristem is the provascular tissue surrounding a centralgroup of pith mother cells. The developmental continuity betweenthe provascular tissue and the mature vascular tissue, and betweenthe pith mother cells and the pith, through transitional stages,indicates that the initial differentiation of vascular tissueand pith takes place in this prestelar tissue. The continuityof vascular differentiation in the area confronting young leavesor incipient leaf positions is interrupted by the formationof leaf gap initials. Developing leaves thus begin to exertinfluence on the vascular system at the prestelar stage. Smallprotoxylem elements with helical cell wall thickening, and distinctiveprotophloem elements are present in the leaf traces, but endabruptly near the junction regions of leaf traces to the meristele.Copyright1994, 1999 Academic Press Initial vascular differentiation, provascular tissue, pith mother cells, shoot apex, Matteuccia struthiopteris     

Characterization of Stelar Initiation in Shoot Apices of Ferns

Procambium is commonly recognized as a vascular meristem inshoot apices of vascular plants. Prestelar tissue comprisingprovascular tissue (PVT) and pith mother cells (PMCs) immediatelysubjacent to the single cell layer of promeristem has been consideredto represent the initial stage of stelar differentiation precedingprocambium and rib meristem in ferns. In addition to characterizationof PVT and PMCs on the basis of cell morphology, cytologicalfeatures and developmental continuity with procambium and ribmeristem, four lines of evidence from studies of shoot apicesof Matteuccia struthiopteris and Osmunda cinnamomea supportthis interpretation of initial differentiation. (1) Differentialstaining by safranin-fast green and crystal violet-erythrosinshows that PVT and PMCs differ in colour reactions from promeristemand resemble procambium and pith meristem, respectively. (2)Comparative ultrastructural study reveals qualitative differencesin the cell membrane system, nuclei, cytoplasm, vacuoles andplastids between promeristem and PVT but similarity of PVT toprocambium. (3) Large droplets of tannins occur in promeristembut not in PVT, PMCs and procambium. (4) Cytochemical studyof the shoot apex of Osmunda shows that carboxylesterase activityis strongly demonstrated in PVT and procambial cells but notin promeristem cells and PMCs. These observations further substantiatethe interpretation that PVT represents initial vascular differentiationand PMCs reflect a commitment to pith development.Copyright1995, 1999 Academic Press Initial vascular differentiation, provascular tissue, differential staining, ultrastructure, tannins, carboxylesterase, shoot apex, Matteuccia struthiopteris, Osmunda cinnamomea     

Effect of Assimilate Supply on Development and Growth Potential of Axillary Buds in Roses

The effect of assimilate supply on axillary bud developmentand subsequent shoot growth was investigated in roses. Differencesin assimilate supply were imposed by differential defoliation.Fresh and dry mass of axillary buds increased with increasedassimilate supply. The growth potential of buds was studiedeither by pruning the parent shoot above the bud, by graftingthe bud or by culturing the bud in vitro. Time until bud breakwas not clearly affected by assimilate supply during bud development,Increase in assimilate supply slightly increased the numberof leaves and leaf primordia in the bud; the number of leavespreceding the flower on the shoot grown from the axillary budsubstantially increased. No difference was found in the numberof leaves preceding the flower on shoots grown from buds attachedto the parent shoot and those from buds grafted on a cutting,indicating that at the moment of release from inhibition thebud meristem became determined to produce a specific numberof leaves and to develop into a flower. Assimilate supply duringaxillary bud development increased the number of pith cells,but the final size of the pith in the subsequent shoot was largelydetermined by cell enlargement, which was dependent on assimilatesupply during shoot growth. Shoot growth after release frominhibition was affected by assimilate supply during axillarybud development only when buds sprouted attached to the parentshoot, indicating that shoot growth is, to a major extent, dependenton the assimilate supply available while growth is taking place.Copyright1994, 1999 Academic Press Assimilate supply, axillary bud, cell number, cell size, defoliation, development, growth potential, meristem programming, pith, Rosa hybrida, rose, shoot growth     

Cytochromes of Mitochondria from Rice Seedlings Germinated under Water and Their Changes during Air Adaptation

Rice seeds were germinated for up to 5 days under water (submerged)and some for another day in air (air-adapted). Control seedswere germinated for 6 days throughout in air. Low-temperaturedifference spectra of shoot mitochondria were compared amongthese three types of seedlings. All cytochromes found in theaerobic seedlings were present in the submerged seedlings. However,there were some differences in the cytochromes b₅₅₃ and c ofthese two types of seedlings. The cytochrome aa₃ peak heightand cytochrome oxidase activity per mitochondrial protein increased1.6- and 2.8-fold, respectively, during air adaptation. Slightlyhigher concentrations of the b-type cytochromes than found inair-adapted mitochondria were already present in submerged mitochondria.The computed difference between the dithionite-reduced differencespectra of mitochondria from submerged seedlings before andafter air adaptation, showed that cytochromes aa₃ and c hadincreased more than cytochrome b₅₅₇ during air adaptation. (Received November 16, 1987; Accepted March 16, 1988)     

Purification and properties of a dissimilatory nitrite reductase of a denitrifying phototrophic bacterium

Nitrite reductase [nitric-oxide : (acceptor) oxidoreductase,EC 1.7.2.1 [EC] ] from a denitrifying phototrophic bacterium, Rhodopseudomonassphaeroides forma sp. denitrificans, was purified. The molecularweight of the enzyme, estimated by gel-filtration, was 80,000.Sodium dodecyl sulfate polyacrylamide gel electrophoresis ofthe purified enzyme showed a single 39,000 molecular weightband, indicating that the enzyme was composed of two subunitsof identical molecular weight. The oxidized form of the enzymeexhibited maximum absorption at 280 nm, 450 nm and 590 nm, andthe reduced form only at 280 nm. The ESR spectrum of a frozensolution of the oxidized enzyme showed a typical spectrum patternof a copper protein, suggesting that two types of Cu²⁺ existedwithin the enzyme. Estimates with an atomic absorption spectrophotometer,revealed two copper atoms per molecule. The optimum pH of theenzyme was 7.0. Km for nitrite was estimated to be 51 µM,and the optimum temperature, 30?C. The enzyme was inhibitedby CO, potassium cyanide and diethyldithiocarbamate and activatedby monoiodoacetate. Phenazine methosulfate, 2,6-dichlorophenolindophenol,horse heart cytochrome c, and cytochrome c₂ from this bacteriumwere suitable electron donors. The enzyme also showed cytochromec oxidase activity. (Received May 4, 1978; )     

Surgical Experiments on the Differentiation of Vascular Tissue in the Shoot Apex of Carrot (Daucus carota L.)

Surgical techniques were applied to the shoot apex of carrot(Daucus carota L.) to test the interpretation that provasculartissue is the initial stage of vascular differentiation andto localize the sources of the influences that control its differentiation.If the apex is isolated laterally by vertical incisions leavingit at the summit of a plug of pith tissue, vascular differentiationproceeds normally and an independent vascular system is formedin the pith plug. If all leaf primordia are systematically suppressed,provascular tissue continues to differentiate as an acropetalextension of the pre-existing vascular system but no furtherdifferentiation occurs. When the apex is isolated laterallyand all leaf primordia are suppressed, provascular tissue continuesto be formed acropetally and is extended basipetally into thepith plug by redifferentiation of pith cells, but no furtherdifferentiation occurs. This tissue reacts positively to histochemicaltests for esterase indicating its vascular nature. If only oneleaf primordium is allowed to develop on an isolated shoot apex,its vascular system develops normally and extends basipetallyinto the pith plug, but there is no extension of provasculartissue into the pith plug. These results support the interpretationthat the initial stage of vascular differentiation is controlledby the apical meristem but that further maturation of vasculartissue depends upon influences from developing leaf primordia.Copyright 2000 Annals of Botany Company Provascular tissue, differentiation, carrot (Daucus carota L.), shoot apex, surgical techniques, leaf primordia     

Respiratory Activities, Cytochrome Composition and Cytochrome c Oxidase Subunit II Levels of Mitochondria from Alloplasmic Wheats Having Aegilops Cytoplasms

Respiratory activities, cytochrome composition and cytochromec oxidase (EC 1.9.3.1 [EC] ) subunit II (COXII) levels of isolatedmitochondria in one euplasmic and four alloplasmic lines ofwheat (Triticum aestivum), having cytoplasms of Aegilops columnaris,Ae.crassa (4x),Ae. mutica and Ae. triuncialis, which show variousgrowth abnormalities, were studied. Cytoplasm-specific variationon the respiratory activities has been revealed among the cytoplasmsof five Triticum and Aegilops species. The cyanide-resistantrespiratory pathway does not seem to contribute to the variationin activity. The low level of state 3 and cytochrome oxidaseactivities were evident in lines having Ae. columnaris and Ae.triuncialis cytoplasms, which also show growth inhibition. Thelow cytochrome oxidase activity in both cytoplasms is consistentwith the low amount of cytochrome aa₃ and the severe reductionin COXII levels. The relatively low amount of cytochrome aa₃in Ae. mutica is also consistent with the reduced level of theCOXII. On the other hand, relatively low amounts of cytochromeb (b-557) are found in the Ae. crassa cytoplasmic line. Theseresults suggest that the growth abnormalities found in the alloplasmicwheat lines are related to the variation in respiratory activitiesand cytochrome composition. In particular, growth inhibitionfound in the alloplasmic wheats having Ae. columnaris and Ae.triuncialis may be caused by the severe depression of respiratoryactivity due to the impaired cytochrome oxidase activity oftheir mitochondria. ³Present address: Department of Biochemistry, Dalhousie University,Halifax, NS, Canada B3H 4H7     

Structural Changes in the Embryonic Axis of Theobroma cacao L., during Germination and Early Seedling Establishment

The changes occurring in the axis of the developing seedlingof Theobroma cacao L. were observed over a 96-h period. It wasfound that during the first 24-h period, greatest change hadoccurred in the hypocotyl; the epicotyl and the root meristemhad apparently remained dormant. In the hypocotyl, vascularizationwhich had begun in the mature embryo continued: starch grainshad disappeared from the cells of the cortex and pith, the latterof which were seen to be binucleate. During the second 24-hperiod, greatest change was seen to have taken place in theroot meristem, resulting in the production of a root which consistedof a central pith, a poly-arch stele, and cortical tissue. Betweenthe root and the hypocotyl an apparently undifferentiated regionwas observed. This region was seen to have differentiated duringthe third 24-h period, leading to the production of a completering of xylem, external to which were groups of phloem, theformer being separated from the latter by parenchymatous tissue.This region is interpreted as being transitional between rootand shoot. During the fourth 24-h period, the adventitious andlateral root primordia initiated earlier, were seen to havedifferentiated to the extent that the disposition of their tissueswas evident.     

The regulation of cytochrome c oxidase of Rhodobacter capsulatus by light and oxygen

In order to distinguish between the regulatory effects of oxygen tension and light intensity on cytochrome c oxidase protein and enzymatic activity cells of Rhodobacter capsulatus were shifted from phototrophic (anaerobic, light) growth to aerobic-light, aerobic-dark and to anaerobic-dark conditions, respectively. During shift-experiments the formation of oxidase protein and regulation of oxidase activity was followed by immunological and enzymatic means. The results support the idea, that the formation of oxidase protein is regulated by oxygen tension and light intensity changes, whereas the regulation of oxidase activity seems only to be correlated to the oxygen tension. A DNA sequence involved in the oxygen-dependent regulation of cytochrome oxidase could be identified in the regulation-deficient oxidase mutant H41 of R. capsulatus. Immunological investigations of cytochrome c ₂ from mutant H41 demonstrated at the same time the participation of the c ₂-polypeptide in the regulation of cytochrome c oxidase.Abbreviations Bchl bacteriochlorophyll - CIE crossed immuno-electrophoresis - DMSO dimethyl sulfoxide     

Relative Contribution of Terminal Oxidases to the Respiration of Isolated Cells

The relative contributions of cytochrome (Cyt) oxidase, cyanide-insensitiveoxidase and residual oxidase to the total respiration of isolatedleaf cells were determined in order to integrate them into amathematical model. During cell incubation, the contributionof cyanide-insensitive oxidase decreased whereas the latentactivity of the enzyme increased. Carbonyl cyanide m-chlorophenyl-hydrazonemainly stimulated the Cyt oxidase activity but did not releasethe latent activity of cyanide-insemitive oxidase. (Received May 10, 1983; Accepted September 21, 1983)     

Succinoxidase and Cytochrome Oxidase in Mitochondria from the Spadix of Arum

1. Mitochondria prepared from young Arum spadix have enoughcytochrome c oxidase to account for the rate at which succinateis oxidized, but succinoxidase activity increases markedly asthe plants mature so that in old material cytochrome oxidaseactivity is only 10 per cent, of succinoxidase. 2. Disintegration of the mitochondria by vibration with ballotini,treatment with digitonin or incubation in the warm reveals anintra-mitochondrial cytochrome c oxidase probably active enoughto account for the fastest rates of succinate oxidation. 3. Succinic dehydrogenase activity is demonstrated and experimentswith p-chloromercuribenzoate indicate that it plays a part inthe oxidation of succinate. 4. Cyanide completely inhibits both external and internal cytochromeoxidase but even at the earliest stages it only reduces succinoxidaseby about 50 per cent. Antimycin A also inhibits succinoxidaseby about 50 per cent.     

Location of the Electron Accepting Site of a b-Type Cytochrome Oxidase in Purified Chromatophore Membranes and Spheroplast Membrane Vesicles from Photosynthetically Grown Rhodopseudomonas sphaeroides

Purified chromatophore membranes and spheroplast membrane vesicleswere prepared from Rhodopseudomonas sphaeroides by the methodof Michels and Konings (1978). The cytochrome c oxidase activityof the spheroplast membrane vesicles was about 15-fold higheron a bacteriochlorophyll basis than that of the chromatophoremembranes. The cytochrome c oxidase activity of the chromatophoremembranes was greatly stimulated (about 10-fold) by an additionof Triton X-100 (0.1%), but there was less stimulated (about1.5-fold) in the case of spheroplasts. The pH optimum of theoxidase activities of the spheroplast membrane vesicles andof chromatophore membranes treated with 0.1% Triton X-100, andthe salt dependencies of the activity of both preparations werethe same. These results show that the membrane-bound b-type cytochromeoxidase of this bacterium accepts electrons from ferrocytochromec on the periplasmic side of the membrane (on the outer surfaceof the spheroplast membrane vesicles). These results also areconsistent with the fact that cytochrome c₂ is located in theperiplasmic space in this bacterium. (Received January 24, 1984; Accepted April 24, 1984)     

7H-Pyridocarbazole, a Probe to Study the Role of Diphosphatidylglycerol in the Functioning of the Cytochrome Oxidase Complex of Mitochondria

Ditercalinium, the generic name for 7H-pyridocarbazole dimer,is a planar aromatic cation which exhibits strong inhibitoryeffects on the respiratory chain of plant mitochondria. By enzymaticassays and spectroscopic studies, the inhibition is shown tooccur at the level of the cytochrome c oxidase complex. It issuggested that the mechanism of inhibition could be due to thecomplexation of the aromatic cation with the diphosphatidylglycerolenvironment essential for cytochrome c oxidase activity ratherthan to a drug-enzyme direct interaction. 7H-pyridocar-bazolecan be used as an appropriate tool to study the role that negativelycharged acidic phospholipids, as diphosphatidylglycerol, playin the functioning of cytochrome c oxidase. (Received July 10, 1989; Accepted February 19, 1990)