TEMPORAL VARIATION IN FORAGING BEHAVIOUR OF PATELLA GRANULARIS (PATELLOGASTROPODA) AND SIPHONARIA CONCINNA (BASOMMATOPHORA) ON A SOUTH AFRICAN SHORE

Foraging activity of two mid- to low- shore species of limpet,Patella granulans (Prosobranchia) and Siphonaria concinna (Pulmonata)from an exposed shore on the Eastern Cape coast of South Africawas monitored. In both species, activity was compared duringspring and neap tides and, in P. granularis between summer andwinter. Rhythms of activity of the two species were similar,with foraging excursions being mainly associated with nocturnallow tide times, although some P. granularis foraged during daytimelow tides. It is suggested that foraging excursions in P. granularisare triggered by wave action. Both species foraged further onspring tides than on neap tides and this is suggested to bea result of the limited time limpets have to forage. P. granulariswas also found to forage further during summer when comparedto winter and the possibility that seasonal micro-algal productivityinfluences foraging distances in limpets is discussed. The foraging activity of both species could be divided into3 distinct phases, a relatively rapid outward phase, a muchslower foraging phase and a rapid homeward phase. Whether ornot these limpets graze throughout an excursion is not known.S. concinna was found to home to a fixed scar, although duringthe experiment some scar-swapping occurred. P. granularis didnot home to a fixed scar but possessed a ‘home range’(approx. 5 cm²) to which it returned after each excursion. Patella granularis was found to move randomly during foraging,whilst S. concinna foraged in a non-random direction -whichtook individuals upshore. No tidal-influence is thought to bepresent in this case and the possibility of a learning componentin the foraging behaviour of certain limpet species in relationto the return to optimal feeding patches is discussed. (Received 18 May 1996; accepted 2 September 1996)     

Rapid separation of carbonic anhydrase isozymes using cellulose acetate membrane electrophoresis

A method is described for the rapid separation of carbonic anhydrase(CA) isozymes by cellulose acetate membrane electrophoresisin which CA activity is detected using the pH-indicating dye,bromcresol purple. This method can detect bovine erythrocyteCA in a 0.3 mm³ sample applied at a concentration of 100 ngcm^–3 (total of 30 pg applied) while at higher concentrationsthree isozymes were observed. It was found, using a potentiometrictechnique, that intact cells of Anabaena flos-aquae (Cyanophyceae)and Chlorella ellipsoidea had no detectable activity while C.saccharophila and Chlamydomonas reinhardtii (Chlorophyceae)had external CA activity. CA activity of the extracts suggestedthe presence of internal CA in all species. After electrophoresisit was found that C. saccharophila and C. reinhardtii had twoisozymes while A. flos-aquae and C. ellipsoidea had only a singledetectable band. Spinach had up to five detectable isozymesthat were difficult to resolve. Incubation of spinach extractwith the CA inhibitor ClO₄^– (500 mol m^–3) inhibitedCA activity by 90% using the potentiometric technique, but afterelectrophoresis had no detectable effect. This technique isuseful in identifying isozymes that are substantially differentin electrical charge and in monitoring CA isozyme activity duringenzyme purification. Key words: Carbonic anhydrase, isozymes, cyanobacteria, microalgae, spinach     

Transient Expression of Exogenous DNA in Intact, Viable Wheat Embryos Following Particle Bombardment

Expression of foreign DNA has been detected in intact, germinatingwheat embryos (Triticum aestivum L.) following bombardment withtungsten particles complexed with a reporter gene encoding thebacterial enzyme ß-glucuronidase (‘GUS’:E.C.3.2.1.31). Expression was detected in situ in individualcells and groups of cells, by supplying the germinating embryoswith the chromogenic substrate of the GUS enzyme, ‘X-gluc’.Expression was dependent on the presence of a constitutive plantpromoter, the Cauliflower Mosaic Virus ‘35S’ promoter,fused to the GUS structural coding sequence. The relative simplicityof this technique recommends its future use for the assay ofregulatory elements which control the spatial and temporal specificityof genes expressed during embryo development. Key words: Transient expression, particle bombardment, wheat embryo, Triticum aestivum L.     

3D spatial point patterns of bioluminescent plankton: a map of the minefield

As the open ocean environment lacks points of refuge from visualpredators, it has favored the evolution of extraordinary adaptationsfor optical concealment, such as vertical migration, transparencyand counterillumination. Bioluminescent plankton, which respondto a mechanical disturbance with a flash of light, are ubiquitousin the ocean and potentially a threat to any organism seekingdarkness as a means to evade detection. Estimating encounterprobabilities for organisms maneuvering through luminescent‘minefields’ requires characterization of the three-dimensionaldistribution patterns of the potential light emitters. In orderto measure nearest neighbor distances (NNDs) of bioluminescentbiota, we have developed a spatial plankton analysis technique(SPLAT) for 3D reconstruction and statistical analysis of thespatial point patterns of identified bioluminescent displays.Analysis of aggregations of bioluminescent biota in the Gulfof Maine (Wilkinson Basin) revealed that encounter probabilitieswere highest in the temperature minimum zone (temperature <5°C)where dinoflagellates (Protoperidineum depressum) exhibitedNNDs of 3.5–3.8 cm in a regular distribution pattern.This technique was also used to examine the internal organizationof thin layers of the bioluminescent copepod, Metridia lucens.Comparison of night-time surface (migrating) and deep (non-migrating)layers indicated complete spatial randomness in both populationsand no significant difference in spacing (NNDs: 9.5–13cm).     

The relationship between community respiration and ETS activity in the ocean

We have studied the relationship between community respiration(R) and enzymatic activity of the electron transport system(ETS) in upper ocean microbial communities (<225 µm)from different oceanic regions. In all except one of the regions,R and ETS were significantly positive correlated. This supportsthe hypothesis that ETS can be widely used to estimate planktonrespiration in natural marine communities (Packard, T.T., Adv.Aquat Microbiol, 3,207–261, 1985). A regression equationwas obtained between all the R and ETS data studied, to deriverespiration from ETS activity. This equation yields a mean errorin the prediction of ±34%, similar to the errors obtainedapplying the equations at each area, but lower than the errorobtained when using the mean R:ETS ratio to determine respiration(±45%). Our results suggest that the use of the ETS-Ralgorithm, along with measurements of ETS activity in seawater,facilitates the estimation of seawater respiratory oxygen consumptionon the mesoscale. This means that by using this approach onecould extend our knowledge of oceanic respiration over largetemporal and spatial scales, and begin to use respiration, notonly productivity, in addressing carbon balance problems inthe upper ocean.     

Short- and long-term movements of the patellid limpet Patella flexuosa within gaps in intertidal mussel beds

Short-term and long-term movements of the patellid limpet Patellaflexuosa Quoy & Gaimard were studied on an intertidal rockyshore where a mytilid, Hormomya mutabilis (Gould), formed anextensive mussel zone. At low tide, all the limpets were found restingwithin gaps in the mussel zone, which were formed after massdislodgement of the mussels by waves. The foraging areas ofall the marked limpets were restricted to within the gaps duringfour 24h diving observations. All of these limpets displayedhoming behaviour after short excursions with a maximum averagedistance of 6.7 cm from their homes. During short-term observationperiods of about 4–5 days, no marked limpets moved fromone gap to another gap across the mussel bed of H. mutabilis.During long-term observation periods of one month, however,10–30% of marked limpets shifted their homes to differentgaps that had a lower limpet density. The shell length of theseemigrants was, in general, significantly smaller than that ofnon-emigrants. The percentage of emigrants tended to be higherin summer and autumn than in winter and spring. The movementof P. flexuosa within patchy habitats is discussed with a focuson the difference in site fidelity between small and large limpets. (Received 14 April 1998; accepted 7 September 1998)     

The Occurrence of 1,3-{beta}-Glucanase in Healthy and Verticillium-infected, Resistant and Susceptible Tomato Plants

Leaf, stem, and root extracts of near-isogenic tomato plantscv. Craigella, resistant and susceptible to Verticillium albo-atrum,showed constitutive 1,3-ß-glucanase activity whichincreased following inoculation with the pathogen. Partiallypurified enzyme extracts were obtained by dialysing a 30–80%ammonium sulphate fraction of the tissue brei. The enzyme hadpH and temperature optima of 5?5 and 44 ?C respectively, withhigh activity between 50 and 60 ?C. The response to laminarinconcentration was linear between 1?2 and 7?5 mg ml^–1.Root inoculation of susceptible plants with 10⁶ propagules ml^–1V. albo-atrum led to a umform 300 per cent increase in all steminternodes except the terminal one, which was 500 per cent ofthe controls. No spatial relationship of enzyme activity tothe localization of fungus within the stem was apparent. Petioles,leaves, and roots of susceptible infected plants similarly showedan increase in activity but less than that in stems. Changedlevels of stern enzyme activity at different times after inoculationwere associated with reductions in the number of vessels containinghyphae. Extracts of plants of the resistant isoline showed increasedglucanase activity over controls, but this was substantiallylower than that in susceptible plants and was associated withthe greatly reduced mycelial colonization in resistant plants. It is concluded that single gene resistance in tomato to Verticilliumis not associated with innately higher levels of 1,3-ß-glucanasein healthy plants. The increased activity in infected plantsis proportional to the overall quantity of pathogen in the plantor of pathogenic metabolites.     

Size-fractionated biomass, photosynthesis and dark CO2 fixation in a tropical oceanic environment

This study examines the spatial distribution and size structureof phytoplankton biomass and productivity in relation to thevertical structrure of the Andaman Sea (northeastern IndianOcean). This region was characterized by low concentrationsof nutrients and high levels of insolation. Nitrogen availabilityappeared to control overall productivity with nitrate-based‘new’ production accounting for 8–24% of thetotal primary production. Euphotic column chlorophyll (chl a)averaged 52.5 mg m^–2 of which a major portion was locatedas a subsurface chl a maximum (SCM) at     

Tagged Mutagenesis and Gene-trap in the Moss, Physcomitrella patens by Shuttle Mutagenesis

The moss, Physcomitrella patens has been used as a useful materialin many fields, because of its simple body plan, ease of genetargeting, and other reasons. Although many mutants have beenreported, no method to isolate the corresponding genes was reported.We developed a gene tagging and gene-trap system in P. patensby using the shuttle mutagenesis technique, which has been usedin the budding yeast. In 5264 tagged lines, 203 mutants withaltered developmental or morphological phenotypes were obtained.In 129 of 4757 gene-trap lines, ß-glucuronidase (GUS)activity was detected in some tissue. Although multiple copiesof a tag were detected in many tagged lines by Southern analyses,most copies are likely integrated at the same locus accordingto PCR analyses.     

Purification, Kinetic and Regulatory Properties of Phosphofructokinase from Chiorella pyrenoidosa

Phosphofructokinase was purified 585-fold from Chlorella pyrenoidosaby using a combination of ammonium sulphate fractionation, filtrationthrough Sepharose 4B and chromatography on DEAE-Sephacel. Enzymestability was maintained by the presence of 50 mM P_i at pH 6.6.The optimum pH for activity was 7.7. Concentrations of substratesrequired to achieve half maximal velocity in the standard assaywere 9 µM (ATP) and 0.2 mM (fructose-6-P). ATP above 0.5mM was inhibitory. Enzyme activity was inhibited by high concentrations(10–100 mM) of P_i but lower concentrations (1–5mM) were effective in relieving the influence of other inhibitorssuch as P enolpyruvate. Inhibition by P-enolpyruvate was greaterat lower pH and with less P_i in reaction mixtures: 50% inhibitioncould be attained with 0.1 mM P-enolpyruvate. Fructose-2,6-bisphosphate,which was shown to be present in Chlorella, had no effect onthe phosphofructokinase. Chlorella appeared to contain onlyone form of phosphofructokinase, possibly in the chloroplast.No pyrophosphate :D-fructose-6-P 1-phospho transferase activitycould be detected. (Received February 20, 1984; Accepted December 5, 1984)     

Fine-scale genetic structure among genetic individuals of the clone-forming monotypic genus Echinosophora koreensis (Fabaceae)

• Background and Aims For rare endemics or endangered plantspecies that reproduce both sexually and vegetatively it iscritical to understand the extent of clonality because assessmentof clonal extent and distribution has important ecological andevolutionary consequences with conservation implications. Asurvey was undertaken to understand clonal effects on fine-scalegenetic structure (FSGS) in two populations (one from a disturbedand the other from an undisturbed locality) of Echinosophorakoreensis, an endangered small shrub belonging to a monotypicgenus in central Korea that reproduces both sexually and vegetativelyvia rhizomes. • Methods Using inter-simple sequence repeats (ISSRs) asgenetic markers, the spatial distribution of individuals wasevaluated using Ripley's L(d)-statistics and quantified thespatial scale of clonal spread and spatial distribution of ISSRgenotypes using spatial autocorrelation analysis techniques(join-count statistics and kinship coefficient, F_ij) for totalsamples and samples excluding clones. • Key Results A high degree of differentiation betweenpopulations was observed (_ST(g) = 0·184, P < 0·001).Ripley's L(d)-statistics revealed a near random distributionof individuals in a disturbed population, whereas significantaggregation of individuals was found in an undisturbed site.The join-count statistics revealed that most clones significantlyaggregate at 6-m interplant distance. The Sp statistic reflectingpatterns of correlograms revealed a strong pattern of FSGS forall four data sets (Sp = 0·072–0·154), butthese patterns were not significantly different from each other.At small interplant distances (2 m), however, jackknifed 95% CIs revealed that the total samples exhibited significantlyhigher F_ij values than the same samples excluding clones. • Conclusion The strong FSGS from genets is consistentwith two biological and ecological traits of E. koreensis: bee-pollinationand limited seed dispersal. Furthermore, potential clone matesover repeated generations would contribute to the observed highF_ij values among genets at short distance. To ensure long-termex situ genetic variability of the endangered E. koreensis,individuals located at distances of 10–12 m should becollected across entire populations of E. koreensis.     

Autotrophic and heterotrophic nanoplankton in the diet of the estuarine copepods Eurytemora affinis and Acartia bifilosa

The ingestion of autotrophic and heterotrophic nanoplanktonby two estuarine copepods, Eurytenora affinis and Acarith bifilosa,was measured in various environmental conditions using the incubationmethod and epifluorescence microscopy. Egg production of thespecies was also deter mined in order to estimate their carbonrequirements. Assuming a gross efficiency of egg productionof 0.3, nanoplanktonic carbon ingested always met the carbonrequirements suggesting that, most of the time, other carbonsources could be unnecessaly. Nanoplankton ingestion by A.bifilosa(from 128 to 1693 cells ind.^–1 h^–1) was dominatedby autotrophic forms (60–97%) and was seriously affectedby high (>100 mg l^–1 suspended particulate matter (SPM)concentrations. Nanoplankton ingestion by E.affinis (from 300to 1049 cells ind^–1 h^–1) was relatively stable incomparison, but this latter species seemed to switch its grazingpressure from autotrophic to heterotrophic forms when SPM concentrationsincreased. Thus, two copepod species, living in the same estuary,presented two different feeding behaviours, probably to maximizeenergy input per unit of energy expenditure. Such differencescould contribute to the spatial and seasonal segregation ofthese species which is usually observed.      

A field study on the physiology of digestion in the Antarctic krill, Euphausia superba, with special regard to chitinolytic enzymes

Endo- and exochitinase activities were determined in the stomachand midgut gland of the Antarctic krill, Euphausia superba.along a transect west of the Antarctic Peninsula. Activitieswere compared with the digestive enzymes protease, cellulase(1,4-ß-D-glucanase) and laminarinase (1,3-ß-D-glucanase)The chlorophyll and protein contents in the surface water ofthe corresponding stations were determined. Enzyme activitieswere characterized by high individual and spatial variations.Chitinolytic activity in the stomach correlated well with alldigestive enzymes investigated. In the midgut gland, a correlationwith cellulase and laminannase was evident. The amount of chlorophylla and phytoplankton protein in the surface water was not correlatedwith enzyme activity. Specific enzyme activity was higher inthe stomach than in the midgut gland. showing individual ratiosfor each enzyme. Elevated endochitinase activity in the stomachsuggests that chitinous food is digested to oligomers in thestomach, while the subsequent degradation to amino sugars occurspredominantly in the midgut gland.     

Poly(A) polymerase in Dormant Embryos of Triticum durum

Triticum durum‘Cappelli’ has a ‘relative’dormancy which can be broken by dry after-ripening at room temperature.The breakage of dormancy in the embryos of T. durum , is accompaniedby a decline in content and a different degree of synthesisof poly(A)⁺RNA. This work studies the activity of poly(A) polymerase(E.C. 2.7.7.19), the enzyme which permits polyadenylation. Anincrease in the activity of this enzyme in parallel with theenhanced rate of germination is revealed. Since poly(A) polymeraseactivity is the same in dormant and non-dormant dry embryos,it seems that the activity of the enzyme is not involved inthe breakage of dormancy. The use of cycloheximide and cordycepinshows the presence of enzymes with different origins: a storedenzyme and one bound to a long lived mRNA, present in dormantand non-dormant embryos, plus an enzyme bound to newly synthesizedmRNA which is mainly active in non-dormant embryos. Since dormancycould be the result of an interaction between hormones, thiswork analyses the effects of GA₃and ABA on poly(A) polymerase.GA₃enhanced poly(A) polymerase activity only in dormant embryoswhile ABA inhibited this activity only in non-dormant embryos.Cycloheximide applied to excised wheat embryos represses thestimulatory and inhibitory effects of GA₃and ABA, respectively.The hormone action on poly(A) polymerase activity is thus dependenton de novo protein synthesis. Results using cordycepin suggestthe presence of a stored mRNA for poly(A) polymerase, togetherwith hormonal regulation of enzyme activity at a translationallevel. Copyright 1999 Annals of Botany Company Triticum durum , wheat, dormancy breakage, poly(A) polymerase, GA₃, ABA, germination.     

Effect of Phosphorus Deficiency on Phosphatase Activity of Cell Walls from Roots of Subterranean Clover

Clover (Trifolium subterraneum L. cv. Mt. Barker) was grownin solution culture with adequate (+P) or no phosphate (–P).Cell walls were extracted from roots in such a way that theywere uncontaminated by other cellular materials. Phosphataseactivity was assayed using p-nitro-phenylphosphate (NPP). Phosphatasebound to cell walls had a pH optimum between 5.0 and 6.0, irrespectiveof the P supply to the plants. Activity of phosphatase boundto cell walls increased with electrolyte concentration of theassay medium at pH 6.5 but not at pH 5.5. This increase in activitywas probably due to a higher degree of ionization of the cellwall at pH 6.5 than at pH 5.5, and to effects of high ionicstrength in decreasing the mutual repulsion of negatively chargedNPP from negative charges on the cell walls. Cell wall-boundphosphatase did not exhibit Michaelis-Menten kinetics: the concentrationof NPP at which activity was half the maximum rate (S_0.5) was0.7 mM for cell walls extracted from roots of both +P and –Pplants. Up to 30% of the phosphatase activity bound to cellwalls could be removed using buffer solutions of high pH andhigh ionic strength which contained Triton X100. Both soluble and cell wall-bound phosphatase(s) of roots increasedin activity with P deficiency. The phosphatase activity of cellwalls increased 1.5 fold as the P concentration in the rootsfell from 0.4–0.2% dry weight. Experiments with sterileroots of clover showed that increases in cell wall-bound phosphataseactivity associated with P deficiency were not due to microbialcontamination. It is argued that phosphatase(s) in cell wallsof roots could make a substantial contribution to the P nutritionof clover in soils deficient in inorganic phosphate by hydrolysingorganic phosphate compounds in the soil. Key words: Phosphatase, Clover, Roots, Phosphorus deficiency, Cell walls     

Plankton community respiration in Bransfield Strait (Antarctic Ocean) during austral spring

Community respiration (R) was determined in Bransfield Straitfrom oxygen changes in water samples incubated in borosilicatebottles maintained at in situ temperature. The respiratory electrontransport system (ETS) activity of seawater communities wasalso measured from the same samples. Both data sets were relatedby the regression equation: log R (mg O₂ m^–3 day^–1)=0.462+0.730xlogETS activity mg O₂ m^–3 day^–1) (r=0.80, n=23). Fromthis equation and 37 ETS activity depth profiles, we calculatedthe integrated (0–100 m) community respiration as beingin the range 1.2–4.5 g O₂ m^–2 day^–1 (mean=2.2).These values do not differ significantly from other publishedresults for the Arctic and Antarctic Oceans. Assuming a respiratoryquotient of unity, the areal respiration ranges between 0.45and 1.69 g C m^–2 day^–1 (mean=0.8). This would representan important sink for the primary production reported for BransStrait. The spatial distribution of community respiration showedhigher values associated with the warmer and phytoplankton-richwaters outflowing from Gerlache Strait into Bransfield Strait,and with the front that separates Bellingshausen Sea watersfrom Weddell Sea waters. We suggest that this pattern of distributionmay be related to the transport of organic matter by the BransfieldCurrent along the front.     

Types, Formation, and Metabolism of Cytokinins in Leaves of Alnus Glutinosa (L.) Gaertn.

The nature of the cytokinins present in mature, expanded leavesof alder (Alnus glutinosa (L.) Gaertn.) was investigated usinga chromatographic system capable of partially resolving zeatinfrom (?)-dihydrozeatin. A zeatin-like cytokinin, present asboth a ‘free’ form and as a polar conjugate, postulatedto the the O-ß-D-glucoside of zeatin, accounted forthe bulk of the cytokinin activity (detected by the soybeancallus assay). Studies on the fate of [8-¹⁴C]zeatin supplied to detached alderleaves indicated this cytokinin to undergo extensive metabolism.An appreciable proportion of the radioactivity was incorporatedinto 80% methanol-insoluble and soluble acidic/neutral fractions,while adenosine- and adenine-like peaks were prominent metabolitesin the basic n-butanol-soluble fraction. Small amounts of glucosides,with the properties of both zeatin-O-ß-D-glucosideand dihydrozeatin-O-ß-D-glucoside were formed, thelatter becoming the most prominent with time. The ability of alder leaves to form glucoside directly from(?)-dihydrozeatin was assessed using the soybean callus assay.(?)-Dihydrozeatin was subject to a relatively rapid, continuousand substantial conversion to its putative O-ß-D-glucosideand cytokinin activity was consequently conserved while, incontrast, leaves supplied with zeatin exhibited a progressiveloss of cytokinin activity and, in agreement with the radioactivityexperiments, produced only a small amount of activity attributableto the putative O-glucosides. The significance of the observed cytokinin metabolism is discussedin relation to the endogenous cytokinin status of the leaf.     

COUPLING MOTOGRAPHIC AND SONOGRAPHIC RECORDING TO ASSESS FORAGING BEHAVIOUR OF PATELLA VULGATA

Motographic recording of locomotor activity and sonographicrecording of rasping were simultaneously obtained in the fieldfrom a group of Patella vulgata L. inhabiting a sheltered shore(Menai Bridge, North Wales). Each limpet was equipped with apiezoelectric transducer, a small amplifier, and a micro-lighterencased in dental acrylic and glued to the shell. A remote filter-amplifier-recorderdevice continuously recorded rasping noise through the activityperiods (nocturnal low tide), when individual movements werealso recorded using an automatic camera. Movements of experimentallimpets did not differ from those of specimens equipped withthe micro-lighter only. Locomotor activity was definitely concentratedduring the first (outgoing) and last (homing) parts of eachactivity phase. Rasping started when limpets remained exposedto air at ebb tide, 1–2 hours before leaving home, andoccurred during the whole period spent away from home. Raspingwas fairly variable in rate, consisting in non-rhythmic boutsseparated by recovery phases. No significant correlation wasobserved between rasping rate and speed variation during differentparts of each excursion. However, the number of rasps performedin each spatial unit of the path was strongly correlated withthe time spent by the limpet in that unit. The data suggestthat in Patella vulgata the spatial organisation of foragingwithin each excursion is based on changes in travel speed relatedto local food density but not on modulation of rasping ratein time. (Received 4 July 1993; accepted 2 August 1993)     

Spatial heterogeneity and genetic variation in the copepod Neocalanus cristatus along two transects in the North Pacific sampled by the Continuous Plankton Recorder

We present a macrogeographic study of spatial heterogeneityin an important subarctic Pacific copepod and describe the firstgenetic analysis of population structure using Continuous PlanktonRecorder (CPR) samples. Samples of Neocalanus cristatus werecollected at a constant depth of 7 m from two CPR tow-routes,(i) an east–west 6500-km transect from Vancouver Island,Canada to Hokkaido Island, Japan, and (ii) a north–southtransect of 2250 km from Anchorage, Alaska to Tacoma, Washington.Analysis of these samples revealed three features of the biologyof N. cristatus. First, N. cristatus undergoes small-scale dielvertical migration that is larger among stages CV–adult(3–6 times more abundant at 7 m at night), than stagesCI–CIV (only 2–4 times higher at night). Secondly,while there were no regions where N. cristatus did not appear,each transect sampled a few large-scale macrogeographic patches.Thirdly, an analysis of molecular variation, using a partialsequence of the N. cristatus cytochrome oxidase I gene, revealedthat 7.3% (P < 0.0001) of the total genetic variation amongN. cristatus sampled from macrogeographic patches by the CPRcould be explained by spatial heterogeneity. We suggest thatspatial heterogeneity at macrogeographic scales may be importantin plankton evolution.     

Purification and Properties of 1-AminocycIopropane-l-carboxylate Synthase of Mesocarp of Cucurbita maxima Duch. Fruits

1-Aminocyclopropane-1-carboxylate (ACC) synthase, which formsAGC from S-adenosylmethionine (SAM), was purified to homogeneityfrom sliced and aged mesocarp tissue of Cucurbita maxima Duch.cv Ebisu fruits, and its enzymatic properties were determined.The specific activity of the purified enzyme was 220 mU/mg proteinat 30°C at 50 µM SAM. Native ACC synthase has a relativemolecular mass of 160 ± 10 kDa and consisted of two subunitsof about 84±3 kDa. S-adenosylhomocysteine (SAH), S-methylmethionine(SMM) and L-methionine did not serve as substrate. The enzymereaction was competitively inhibited by aminoethoxyvinylglycine(AVG) (Ki, 2.5 µM), aminooxyacetic acid (Ki, 40 µM)and SAH (Ki, 30 µM). The reaction was also strongly inhibitedby semicarbazide, and less effectively by homocysteine. Theenzyme was rapidly inactivated by its substrate, SAM in thepresence of pyridoxalphosphate (PLP), but in the absence ofPLP, SAM-induced inactivation was much slower. Inactivationdid not occur by SAH and SMM, SAM analogs without substrateactivity. Pyridoxal phosphate was an essential cofactor to beadded to a reaction mixture for maximum activity, but an enzymepreparation from which pyridoxal phosphate was removed by SephadexG-25 gel filtration exhibited one-eighth activity which wasinhibited by semicarbazide, this indicating that a small amountof pyridoxal phosphate is firmly bound to the enzyme. (Received May 6, 1986; Accepted May 20, 1986)