pH and Temperature Dependence of Tyrosine Z Decay Kinetics in Tris-Treated PSII Particles Studied by Time-Resolved EPR

The decay kinetics of tyrosine Z (Y_z) in Tris-treated PhotosystemII particles were measured by time-resolved EPR at differentpH values (pH 5.5 to 7.5) between 230 and 297 K. Y_z inducedby laser flashes decayed in a biphasic wav; t values were about100 ms in the fast phase and about 1 s in the slow one, respectively,at room temperature. The fast phase was attributed to a recombinationof charges on Y_z and Q^–_A. The activation energies forthe reaction of Y_z with Q^–_A between 245 and 297 K havebeen estimated to be more than 38 kJ mol^–1 at pH (>6.5)and less than 32 kJ mol^–1 at pH (<6.0), respectively.The activation energy gap between high pH (>6.5) and lowpH (<6.0) ranges was found to be E=4.4 kJ mol^–1. Judgingfrom the fact that pH 6.2 appears to be the border between thehigh and low pH regions, we suggest that the kinetics of Y_zis strongly influenced by the dissociation of the nearby histidineresidue. (Received February 21, 1997; Accepted June 24, 1997)     

Diurnal mixed layers and the long-term dominance of Microcystis aeruginosa

The population dynamics of the cyanobacterium Microcystis aeruginosain a hypertrophic, subtropical lake (Hartbeespoort Dam, SouthAfrica) were followed over 4.5 years. We examined the hypothesisthat M.aeruginosa dominated (>80% by volume) the phytoplanktonpopulation up to 10 months of each year because it maintaineditself within shallow diurnal mixed layers and was thus ensuredaccess to light, the major limiting resource. Wind speeds overHartbeespoort Dam were strong enough to mix the entire epilimnionthrough Langmuir circulations only 12% of the time. At othertimes solar heating led to the formation of diurnal mixed layers(z₁) that were shallower (<2 m) than the euphotic zone (z_cu;mean = 3.5 m, range: 0.45–8.4 m) while the seasonal mixedlayer (z_m) was always deeper than z_cu (range: 7–18 m).By means of its buoyancy mechanism M.aeruginosa maintained thebulk of its population within z₁, while non-buoyant speciessank into dark layers. Adaptation to strong light intensitywas implicated from low cellular chlorophyll a content (0.132µg/106 cells) and high I_k (up to 1230 µE m^–2s^–1). Ensured access to light, the post-maximum summerpopulations persisted throughout autumn and winter, despitesuboptimal temperatures, by sustaining low losses. Increasedsedimentation losses caused a sharp decline of the populationat the end of winter each year, and a short (2–3 months)successional episode followed, but by late spring M.aeruginosawas again dominant. The data from Hartbeespoort Dam point outthe importance of distinguishing between z_m and z₁, and showthe profound effect that the daily pattern of z₁ as opposedto the seasonal pattern of z_m, can have on phytoplankton populationspecies composition.     

Autolysis Promotes the Extension Capacity of Zea mays Coleoptile Cell Walls in Response to Acid pH Solutions

The relationship between autolytic degradation of ß(1–3),(1–4)-D-glucanand acid pH-induced extension of isolated Zea mays cell wallshas been investigated using a constant-load extension technique.Acidic buffer (4.5) was able to induce an additional extension(E_a) on cell walls already extended at pH 6.8 buffer under a20 g-mass load, indicating that the additional extension (E_a)was the parameter that better represented the effect of thedifferent treatments on the mechanical properties of maize coleoptilecell walls. The additional extension in response to acidic pHwas higher when cell walls had been previously autolysed for24 h at pH 5.5. Furthermore, the acid-pH effect was dependenton the presence during the constant load extension of some thermo-labilefactors, suggesting the participation of expansins. Acid pHincreased E_a of native cell walls through an increase in theplastic extension (E_p) in agreement with a one step mechanismleading directly to irreversible (plastic) wall extension assuggested by Cosgrove (1977). The autolytic degradation of ß(1–3),(1–4)-D-glucan was also able to modify the mechanicalproperties of maize coleoptile cell walls increasing its elasticextension (E_e) in response to pH 4.5 buffer but that modificationonly leads to an increase in wall extension when expansins areactive, suggesting a cooperation between ß-glucanturnover and expansin action. (Received August 5, 1998; Accepted March 16, 1999)     

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     

Comparison of carbon-specific growth rates and rates of cellular increase of phytoplankton in large limnetic enclosures

Potential carbon-specific growth rates of phytoplankton wereestimated from a series of measurements of photosynthetic radio-carbonuptake over 4- and 24-h exposure periods in the light fieldsof three large limnetic enclosures (‘Lund Tubes’),each providing different limnological and trophic conditions.Photosynthetic behaviour and short-term, chlorophyll-specificcarbon-fixation rates conformed to well-established criteriabut, over 24 h, the net retention represented 23–82% ofthe carbon fixed during the daylight hours. Potential mean growthrates (k'_p, of the photo-autotrophic community were calculatedas the net exponential rates of daily carbon-accumulation relativeto derived, instantaneous estimates of the cell carbon-content.Apparent actual community growth rates (k'_D were calculatedas the sum of the exponential rates of change of each of themajor species present, corrected for probable rates of in situgrazing and sinking, and expressed relative to the fractionof total biomass for which they accounted. The correspondingvalues were only occasionally similar, k'_p generally exceedingK'_D by a factor of between 1 and 30 or 1 and 14, depending uponthe carbon:chlorophyll ratio used. The ratio, K'_p/K'_D was foundto vary inversely both to k'_D and to k_n, the net rate of changein phytoplankton biomass, suggesting that measured carbon fixationrates merely represent a capacity for cellular increase which,owing to other likely limitations upon growth, is seldom realized.Apparent rates of loss of whole cells do not account for theloss of carbon; that the ‘unaccounted’ loss rates(K'_p–K'_D varied in direct proportion to K'_p (i.e., losseswere least when chlorophyll-specific photosynthetic productivitywas itself limited) is best explained by physiological voidingof excess carbon (for instance, by respiration, photorespiration,excretion) prior to the formation of new cells.     

Regulation of the Cytoplasmic pH of Chara corallina: Response to Changes in External pH

Effects of external pH (pH_o) on the cytoplasmic pH (pH_c) ofChara corallina have been measured with the weak acid 5, 5-dimethyloxazolidine-2,4-dione (DMO) following standardized pretreatment of cells insolutions at pHo 4.5, 6.3 and 8.3. Irrespective of pH_c duringpretreatment, pH_o responded to pHo during the experimental periodsof 150–180 min or (in one experiment) 90–110 min.There were increases or decreases of about 0.5 in pHo when cellswere transferred from pH_o 4.5 to 8.3 or vice versa. In the darkpH_c was 0.2–0.3 units lower than the corresponding valuein the light. The results are discussed in relation to the factorsinvolved in the regulation of pH_c in C. corallina, which maybegin to break down below about pH_o4.5, as indicated by relativelylarge decreases in pH_c at low pH_o. Key words: Chara corallina, Cytoplasmic pH, External pH, DMO     

The Effects of pH on the Ionic and Electrical Properties of the Internodal Cells of Chara australis

The effects of pH on the resting and action potentials and onthe fluxes of potassium, sodium, and chloride across the membranesof internodal cells of Chara australis have been investigated. Experiments were carried out in an artificial pond water (A.P.W.)of standard composition: CaCl₂, 01 mM; KCl, 0.1 mM; NaCl, 1.0mM. The resting potential decreased as the pH was lowered from6.5, being depolarized by about 75 mV at pH 4.5. Measurementsof the ion fluxes as a function of pH suggested that this depolarizationwas caused by an increase in the permeability to sodium anda decrease in permeability to potassium at pH 4.5. Action potentialsof constant peak value can be elicited for some time at pH 4.5,but after 20 min or so the cell becomes refractory. All theseeffects on resting and action potentials are fully reversible.We briefly speculate about the mechanism of these pH effects.     

Two Polypeptides Inducible by Low Levels of CO2 in Soluble Protein Fractions from Chlorella regularis Grown at Low or High pH

Previous studies suggested that certain protein(s) other thancarbonic anhydrase might play an important role in the facilitatedtransport of dissolved inorganic carbon (DIC) from the mediumto the site of CO₂ fixation by ribulose-1,5-bisphosphate carboxylase/oxygenasein the unicellular green alga Chlorella regularis adapted tolow-CO₂ (ordinary air) conditions [Shiraiwa et al. (1991) Jpn.J. Phycol. 39: 355; Satoh and Shiraiwa (1992) Research in Photosynthesis,Vol. III, p. 779]. The proteins that might be involved in thisfacilitated transport of DIC were investigated by pulse-labelingof induced proteins with ³⁵S-sulfate during adaptation of cellsgrown under high-CO₂ conditions to low CO₂. Analysis by SDS-PAGErevealed that synthesis of two polypeptides, with molecularmasses of 98 and 24 kDa, respectively, was induced under low-CO₂conditions. The 24-kDa polypeptide was induced at pH 5.5 butnot at pH 8.0, whereas the 98-kDa polypeptide was induced atboth pH 5.5 and pH 8.0. The possible role of these polypeptidesin the facilitated transport of DIC in Chlorella regularis isdiscussed. (Received October 30, 1995; Accepted February 26, 1996)     

Joint field experiments for comparisons of measuring methods of photosynthetic production

During the 1st GAP Workshop at Konstanz in April 1982 comparativemeasurements of phytoplankton primary production by severaltechniques were conducted simultaneously at an offshore stationin Lake Konstanz and an experimental algal pond. Suspended glassbottle exposure techniques using ¹⁴C and ¹³C uptake gave P_z(mg C m^–3 h^–1) values which varied considerablynear-surface, but estimates of areal rates for the euphoticzone P_cu(mg C m^–3 h^–1) which were reasonably close.In the lake, P_z, from a vertical tube exposure (with ¹⁴C uptake)was greater than rates derived for integrated bottle samples.The oxygen bottle method permitted a good estimate of compensationdepth, corresponding to in situ growth studies. There were difficultiesin direct comparison between O₂ and carbon methods. Correlationbetween them for P_z was good in the lake but poor in the pond,both for suspended bottle and vertical tube methods. This seriesdemonstrates that despite reasonable overall estimates, comparativelyminor methodological differences in experimental technique cancause large variation. ⁺ Coordinator of the group for comparative measurements of photosyntheticproduction at the GAP Workshop, Konstanz, April 1982. ^*This paper is the result of a study made at the Group for AquaticPrimary Productivity (GAP) First International Workshop heldat the Limnological Institute, University of Konstanz, in April1982.     

Mechanisms of the acido- and thermo-phily of Cyanidium caldarium Geitler I. Effects of temperature, pH and light intensity on the photosynthetic oxygen evolution of intact and treated cells

Photosynthetic oxygen evolution in an acido- and thermo-philicunicellular alga, Cyanidium caldarium, was measured under variousconditions, using a Clark-type oxygen electrode. 1). Maximum Hill reaction activity with p-benzoquinone as theHill oxidant was obtained at 45°C in a wide pH range from1.0 to 7.0. 2) The pH activity curve showed two peaks at pH3.0 and 7.0. The Hill activity had an optimum at pH 3.0 in cellspreilluminated under strong light (300,000 lux, 30 min, 40°C).Sonication of algal cells abolished the pH 3.0 component ofthe Hill reaction producing an activity maximum at pH 7.0. 3)Endogenous O₂ evolution in the absence of the Hill oxidant,which lasted for several minutes after illumination, had a maximumat pH 7.0. 4) This endogenous O₂ evolution was abolished bysonication. 5) KCN inhibited endogenous O₂ evolution, but notthe Hill reaction in the presence of p-benzoquinone. (Received August 19, 1974; )     

Evidence for Several Galactan Synthases in Flax (Linum usitassimum L.) Suspension-Cultured Cells

A membrane fraction from flax cells was able to incorporate[¹⁴C]galactose from UDP-D-[¹⁴C]galactose in vitro. The productsof the reaction, characterized by methylation analysis, consistedof a rß-1,4-galactan (solubilized mainly in water)and a rß-1,3- rß-1,6-galactan (solubilizedmainly in alkali). These results indicated the presence of severalgalactan synthase complexes, as did a profile of the relationshipbetween pH and activity which revealed both a maximum at pH6.5 and a shoulder at pH 8. Moreover, galactan synthase activitieswere found at two densities: 1.125 g cm^–3 (Golgi membranes)and 1.07–1.08 g cm^–3 (corresponding to low-densityvesicles). Partial characterization of one enzymatic system (maximaly activeat pH 8 in the presence of 5 mM MgCl₂) was achieved. The K_mfor UDP-galactose and V_max were 38 µM and 4.5 nmol h^–1(mg protein)^–1, respectively. (Received June 6, 1993; Accepted September 22, 1993)     

Quantitative Analysis of ATP-Dependent H+ Efflux and Pump Current Driven by an Electrogenic Pump in Nitellopsis obtusa

Internodal cells of Nitellopsis were made tonoplast-free byperfusion with a medium containing EGTA. Cytoplasmic concentrationsof solutes were controlled by a second perfusion with mediaof known composition. The electrogenic pump current (I_p), whichwas calculated from electrical data obtained from cells withand without ATP, was compared with the current carried by H⁺(I_H⁺) across the plasma membrane. A close correlation betweenI_p and I_H⁺ was found under various internal and external conditions.(1) I_p and I_H⁺ depended on the internal ATP and showed Michaelis-Mententype saturation curves. For I_p, K_m was 120 µM and themaximum current V_max was 15.1 mA m^–2, while for I_H⁺, K_mwas 160 µM and V_max was 16.6 mA m^–2. (2) I_p andI_H⁺ showed almost the same I_H²⁺ dependence. The Mg²⁺-dependentI_p was 19.5 mA m^–2, while the Mg²⁺-dependent I_H²⁺ was17.7 mA m^–2. (3) I_H²⁺ was maximal at an external pH of8 and decreased both in acidic and alkaline pH ranges. I_p wasnearly equal to I_H⁺ in the pH range between 8 and 5. (4) I_H⁺became maximal at an internal pH of 7.3, which is nearly thesame as the pH for maximal electrogenecity found by Mimura andTazawa (1984). All these facts support the idea proposed in our previous paper(Takeshige et al. 1985) that the electrogenic ion pump locatedin the plasma membrane of Nitellopsis is the H⁺ pump. ¹ Dedicated to Professor Dr. Erwin Bünning on the occasionof his 80th birthday. (Received June 21, 1985; Accepted December 20, 1985)     

Cytosolic pH regulates GCl through control of phosphorylation states of CFTR

Our objective inthis study was to determine the effect of changes in luminal andcytoplasmic pH on cystic fibrosis transmembrane regulator (CFTR)Cl conductance(G_Cl). Wemonitored CFTRG_Cl in the apicalmembranes of sweat ducts as reflected byCl diffusion potentials(V_Cl) andtransepithelial conductance(G_Cl). We foundthat luminal pH (5.0-8.5) had little effect on thecAMP/ATP-activated CFTRG_Cl, showing thatCFTR G_Cl ismaintained over a broad range of extracellular pH in which it functionsphysiologically. However, we found that phosphorylation activation ofCFTR G_Cl issensitive to intracellular pH. That is, in the presence of cAMP and ATP [adenosine5'-O-(3-thiotriphosphate)],CFTR could be phosphorylated at physiological pH (6.8) but not at lowpH (~5.5). On the other hand, basic pH prevented endogenousphosphatase(s) from dephosphorylating CFTR.After phosphorylationof CFTR with cAMP and ATP, CFTRG_Cl is normallydeactivated within 1 min after cAMP is removed, even in the presence of5 mM ATP. This deactivation was due to an increase in endogenousphosphatase activity relative to kinase activity, since it was reversedby the reapplication of ATP and cAMP. However, increasing cytoplasmicpH significantly delayed the deactivation of CFTRG_Cl in adose-dependent manner, indicating inhibition of dephosphorylation. Weconclude that CFTRG_Cl may beregulated via shifts in cytoplasmic pH that mediate reciprocal controlof endogenous kinase and phosphatase activities. Luminal pH probably has little direct effect on these mechanisms. This regulation of CFTRmay be important in shifting electrolyte transport in the duct fromconductive to nonconductive modes.

     

Force treadmill for measuring vertical and horizontal ground reaction forces

Weconstructed a force treadmill to measure the vertical, horizontal andlateral components of the ground-reaction forces (F_z,F_y,F_x, respectively) and the ground-reaction force moments(M_z,M_y,M_x), respectively exerted bywalking and running humans. The chassis of a custom-built, lightweight(90 kg), mechanically stiff treadmill was supported along its length bya large commercial force platform. The natural frequencies of vibrationwere >178 Hz for F_z and >87Hz for F_y, i.e., well above thesignal content of these ground-reaction forces. Mechanical tests andcomparisons with data obtained from a force platform runway indicatedthat the force treadmill recordedF_z,F_y,M_x andM_y ground-reaction forces andmoments accurately. Although the lowest natural frequency of vibrationwas 88 Hz for F_x, thesignal-to-noise ratios for F_x andM_z were unacceptable. This devicegreatly decreases the time and laboratory space required for locomotionexperiments and clinical evaluations. The modular design allows forindependent use of both treadmill and force platform.

     

A carrier-mediated mechanism for pyridoxine uptake by human intestinal epithelial Caco-2 cells: regulation by a PKA-mediated pathway

Vitamin B₆ is essential for cellular functions and growth due to its involvement in important metabolic reactions. Humans and other mammals cannot synthesize vitamin B₆ and thus must obtain this micronutrient from exogenous sources via intestinal absorption. The intestine, therefore, plays a central role in maintaining and regulating normal vitamin B₆ homeostasis. Due to the water-soluble nature of vitamin B₆ and the demonstration that transport of other water-soluble vitamins in intestinal epithelial cells involves specialized carrier-mediated mechanisms, we hypothesized that transport of vitamin B₆ in these cells is also carrier mediated in nature. To test this hypothesis, we examined pyridoxine transport in a model system for human enterocytes, the human-derived intestinal epithelial Caco-2 cells. The results showed pyridoxine uptake to be 1) linear with time for up to 10 min of incubation and to occur with minimal metabolic alteration in the transported substrate, 2) temperature and energy dependent but Na⁺ independent, 3) pH dependent with higher uptake at acidic compared with alkaline pHs, 4) saturable as a function of concentration (at buffer pH 5.5 but not 7.4) with an apparent Michaelis-Menten constant (K_m) of 11.99 ± 1.41 µM and a maximal velocity (V_max) of 67.63 ± 3.87 pmol · mg protein^-1 · 3 min^-1, 5) inhibited by pyridoxine structural analogs (at buffer pH 5.5 but not 7.4) but not by unrelated compounds, and 6) inhibited in a competitive manner by amiloride with an apparent inhibitor constant (K_i) of 0.39 mM. We also examined the possible regulation of pyridoxine uptake by specific intracellular regulatory pathways. The results showed that whereas modulators of PKC, Ca⁺²/calmodulin (CaM), and nitric oxide (NO)-mediated pathways had no effect on pyridoxine uptake, modulators of PKA-mediated pathway were found to cause significant reduction in pyridoxine uptake. This reduction was mediated via a significant inhibition in the V_max, but not the apparent K_m, of the pyridoxine uptake process. These results demonstrate, for the first time, the involvement of a specialized carrier-mediated mechanism for pyridoxine uptake by intestinal epithelial cells. This system is pH dependent and amiloride sensitive and appears to be under the regulation of an intracellular PKA-mediated pathway. vitamin B₆; intestinal transport; transport regulation; Caco-2 cell     

Photoinhibition and the diurnal variation of phytoplankton photosynthesis--I. Development of a photosynthesis--irradiance model from studies of in situ responses

Diurnal series of fluorescence and photosynthesis assays wereconducted in high altitude (3803 m), tropical (16°), LakeTiticaca (Peru/Bolivia). Near-surface diurnal thermoclines formedon typical days of high photon flux density (PFD, {small tilde}2000 µE m^–2 s^–1). In the depth range of diurnalstratification profiles of in vivo fluorescence, both without(F_a and with (F_b DCMU, exhibited a mean decrease of 64% frommorning to mid-day, but little change (mean increase of 1.5%)through the afternoon. Three times during the day surface, mid-depth(3–5 m) and deep (15–20 m) phytoplankton sampleswere incubated with H¹⁴CO₃^– under short (<2 h) exposuresto a range of in situ PFDs. Comparison of phytoplankton in differentsamples (ANOVA) showed identical photosynthetic response insunrise (isothermal) samples but a significant drop in surfaceand mid-depth photosynthesis at all PFDs during times of diurnalstratification. Similarly, both low-light () and light-saturated(P² _max photosynthetic parameters were lower in mid-day surfacesamples compared to deep samples. In addition, previously photoinhibitedsamples had a higher threshold intensity for photoinhibition,I_T. These results, together with diurnal time series of fluorescencefrom in situ incubations, demonstrate that recovery from extendedepisodes of photoinhibition during diurnal stratification isslower than suggested by previous observations in vitro. Photosynthesisby near-surface phytoplankton is different in light increasingup to I_T than light decreasing from I_T. This effect can be modeledby reducing and P_max as a function of the maximum photoinhibitingPFD in the diurnal light history. ¹Present address: Division of Molecular Plant Biology, Universityof California, Berkeley, Berkeley, CA 94720, USA     

Purification and characterization of the enzymes of fructan biosynthesis in tubers of Helianthus tuberosus 'Colombia': I. Fructan: fructan fructosyl transferase

Fructan: fructan fructosyl transferase (FFT), one of the enzymesinvolved in the synthesis of ß-2,1 linked fructosepolymers has been purified 205-fold from tubers of Helianthustuberosus harvested in the accumulation phase. The molecularweight of the native as well as the SDS-denatured protein isapproximately 70 kDa. On IEF, the protein was separated intofive molecular species with pl values between pH 4.5–5.0.The optimum pH for fructosyl transfer activity was between 5.5–7.0.Temperature optimum was in the range of 25-35° C; the Q10value between 25 and 5° C was 1.14. FTT catalysed the self-transferof fructosyl groups with GF₂, GF₃, GF₄ or GF₅ as substrate andacceptor. The rate of elf-transfer with both GF₂ and GF₃ increasedlinearly with substrate concentration up to 100 mol m^–3and was still not saturated at 600 and 300 mol m^–3, respectively.FFT was unable to hydrolyse GF or to catalyse the self-transferwith GF but could mediate the transfer of fructosyl units frominulin on to GF. Key words: Fructan: fructan fructosyl transferase, Helianthus tuberosus, Jerusalem artichoke, purification, kinetics     

Changes in lower limb volume in humans during parabolic flight

Variations in gravity [head-to-footacceleration (G_z)] inducehemodynamic alterations as a consequence of changes in hydrostatic pressure gradients. To estimate the contribution of the lower limbs toblood pooling or shifting during the different gravity phases of aparabolic flight, we measured instantaneous thigh and calf girths byusing strain-gauge plethysmography in five healthy volunteers. Fromthese circumferential measurements, segmental leg volumes werecalculated at 1, 1.7, and 0 G_z.During hypergravity, leg segment volumes increased by 0.9% for thethigh (P < 0.001) and 0.5% for thecalf (P < 0.001) relative to1-G_z conditions. After suddenexposure to microgravity following hypergravity, leg segment volumeswere reduced by 3.5% for the thigh (P < 0.001) and 2.5% for the calf (P < 0.001) relative to 1.7-G_zconditions. Changes were more pronounced at the upper part of the leg.Extrapolation to the whole lower limb yielded an estimated 60-mlincrease in leg volume at the end of the hypergravity phase and asubsequent 225-ml decrease during microgravity. Although quantitativelyless than previous estimations, these blood shifts may participate inthe hemodynamic alterations observed during hypergravity and weightlessness.

     

Effects of Ammonia and Methylamine on Cl- Transport and on the pH Changes and Circulating Electric Currents Associated with HCO3- Assimilation in Chara corallina

Low concentrations of ammonia and methylamine greatly increaseCl^– influx into Chara corallina. Both amines have theirmaximum effect at pH 6.5–7.5. The amine stimulation ofCl^– influx is small below about pH 5.5. Above pH 8.5 theremay be inhibition of influx by amines. Concentrations of 10–25µM ammonia are sufficient to cause the maximum stimulationof Cl^– influx; the corresponding methylamine concentrationsare 0.1–0.2 mM. It is concluded that entry of amine cations(NH₄^$ and CH₃NH₃^$), rather than unionized bases (NH₃ and CH₃NH₂),causes Cl^– transport to be increased. Increases in rates of Cl^– transport are not necessarilyaccompanied by effects on HCO₃^$ assimilation and OH^– efflux.Measurements of localized pH differences at the cell surfaceand of circulating electric currents in the bathing solutionshow that these phenomena are only significantly affected byammonia at or above 50 µM and by methylamine at or above1.0 mM. The significance of the effects of amines is assessedin relation to current ideas about transport of Cl^–, HCO₃^–,and OH^–.     

Studies on the phosphomannose isomerase of Amorphophallus konjac C. Koch I. Its isolation and some enzymic properties

A method is described for isolating phosphomannose isomerasefrom young konjak corms. The enzyme is believed to catalyzethe mannose forming reaction in growing corm tissues. The purifiedenzyme preparation was free from phosphoglucose isomerase activity,and was stable at pH 6–9. Maximum enzyme activity wasobserved at pH 6.5–7.0. The molecular weight of the enzymewas estimated as 45,000 using Sephadex gel filtration. The followingkinetic parameters were obtained: Km (mannose-6-P), 0.73 mM,K_eq (fructose-6-P/mannose-6-P), 1.06 at pH 6.5, and activationenergy, 11,600 cal/mole. The enzyme was inhibited by the metalbinding agents EDTA, o-phenanthroline, and a,a'-bipyridyl. Theinhibitory effect of these agents was markedly influenced bythe pH level of the incubation mixture, being more pronouncedat pH 6 than at pH 8. ¹ This paper constitutes part 3 of studies on konjak mannanbiosynthesis. (Received March 3, 1975; )