Purification and characterization of cyclic 3′5′-nucleotide phosphodiesterase D3 from Sinorhizobium fredii MAR-1

The cyclic 35-nucleotide phosphodiesterase D3 was purified from Sinorhizobium fredii MAR-1. The native enzyme had a molecular weight of approximately 44.5kDa and a subunit molecular weight of approximately 21kDa as judged by SDS-gel electrophoresis. The pH optimum of the enzyme for the hydrolysis of cyclic AMP was approximately 6.0 with both acetate and Tris-maleate buffers. The optimum temperature for hydrolysing cyclic AMP was approximately 50C. No metal ion was required for activity and EDTA up to 2.5mM did not markedly affect the enzyme. However, methylxanthines, adenine and adenosine as well as 5-AMP, ATP, ADP and metal ions like Zn²⁺, Fe²⁺, Pb²⁺, Al³⁺ and Fe³⁺, were strongly inhibitory at 2.5mM.The D3 enzyme could hydrolyse both cyclic AMP and cyclic GMP with the apparent K _m for cyclic AMP of approximately 0.23M.     

Mode of uptake of sterol by Arthrobacter simplex

The mechanism of uptake of water-insoluble -sitosterol by a newly isolated strain of Arthrobacter simplex SS-7 was studied. The production of an extracellular sterol-pseudosolubilizing protein during growth of A. simplex on -sitosterol was demonstrated by isolating the factor from the cell-free supernatant and its subsequent purification by Sephadex G-150 column chromatography. The M _r of the purified sterol-pseudosolubilizing protein determined by SDS–PAGE was 19kDa. The rate of sterol pseudosolubilization (5.2×10^–3g l^–1h^–1) could not adequately account for the rate of sterol uptake (72×10^–3g l^–1h^–1) and the specific growth rate (56×10^–3 h^–1). However in the unfavourable growth condition, when the cells were treated with sodium azide at the level of 30–60% of MIC, the sterol pseudosolubilization accounted for nearly 74% of the total growth containing 96% free cells. Cellular adherence to substrate particles was found to play an active role in the normal growth of the strain on -sitosterol. Unlike sodium acetate-grown cells, whose surface activity was negligible (60mNm^–1), the sterol-grown cells had strong surface activity (40mNm^–1). The high lipid content and long chain fatty acids in the cell-wall of -sitosterol-grown cells probably contribute to the high sterol adherence activity of the cells.     

Probable male heterogamety in the deep-sea fish Bathylagus wesethi (Teleostei: Bathylagidae)

A presumed XY chromosome pair is described fromt estis squashes from the mesopelagic deep-sea fish Bathylagus wesethi, whose 2N chromosome number was determined as 34-XY. Although the metacentric X-chromosome is the largest in the entire compliment, the Y is the smallest and only acrocentric element. The positive heteropycnosis of the sex elements was not easily distinguishable in the nuclei of first meiotic prophase. Tetraploid nuclei were observed in peripheral supporting cells of the testis. Males of at least two other congeners have similar karyotypes.     

Unique sequences for two HLA-DRB1 genes expressed on distinct DRw6 haplotypes

We have used restriction fragment length polymorphism (RFLP) analysis and DNA sequencing to characterize two distinct DRB1 alleles expressed on DRw52 and DQw7-associated haplotypes but not readily defined by conventional DR serology. These two haplotypes, designated HLA-D HAG and PEV, react variably with DRw13(w6), DRw14(w6), and the more broad DR 3+6 antisera. Analysis of RFLP revealed that HLA-D HAG and PEV are associated with different DRw52 variants, and that HAG is indistinguishable from DRw18(3) haplotypes. Sequencing of the HAG and PEV DRB1 genes showed each to represent novel alleles. Nevertheless, these sequences show similarities with the other alleles of the DR5, w6, and w8 family. HAG (DRB1*1303) appears to have arisen either from two recombinational events involving at least three DRB1 sequences (DRB1*1101, DRB1*0803, DRB1*0401) or from a single recombinational event together with multiple point mutational events. PEV appears to represent a DRB1*1301-1302/DRB1*1101 recombinant allele, with recombination having occured in the region of bases 175 – 198. The results of this study suggest that the DRw52 family haplotypes is derived from a relatively restricted number of ancestral sequences, with diversity among DRB1 alleles within this family arising through gene conversion or recombination events.     

Plant regeneration via somatic embryogenesis in Styrian pumpkin: cytological and biochemical investigations

Somatic embryo formation was induced from cotyledon explants of Styrian pumpkin (Cucurbita pepo L. subsp. pepo var. styriaca Greb.) by using a solid MS medium supplemented with 16.11M NAA and 4.44M BA or 26.85M NAA and 13.32M BA. The callus proliferation was more efficient on medium supplemented with 26.85M NAA and 13.32M BA. In contrast, the embryogenic response was higher on medium with lower concentrations of growth regulators (16.11M NAA and 4.44M BA). The time needed for embryo induction did not depend on medium composition. Embryos in globular stage were transferred to three different maturation media, containing 2.89M GA₃ in combination with 0.54M NAA, 11.42M IAA and growth regulator-free medium. The germination rate was the highest when embryos were cultured on medium with 11.42M IAA. Plantlets grown on this medium achieved maturity suitable for transplantation into soil within 9 to 10weeks. The regenerated plants were successfully transferred into field and developed fertile flowers and set fruits. Biochemical analysis showed significant lower total glutathione levels among in vitro grown plantlets compared to seedlings grown in soil. When the plantlets were transferred into soil, they reached a normal size within a month and the glutathione concentration was comparable to seed-derived plants at the same developmental stage. Transmission electron microscopy was used to investigate possible differences in the ultrastructure of cells from callus cultures, and leaf cells of regenerated and seed-derived plants. Differences in the ultrastructure were found within chloroplasts which contained only single thylakoids, large starch grains and small plastoglobuli in callus cells in comparison to leaf cells, which possessed a well developed thylakoid system, small starch grains and large plastoglobuli.     

The contribution of size-fractionated plankton to biomass and primary production of phytoplankton in saline–alkaline ponds

Primary productivity, biomass and chlorophyll-a of size fractionated phytoplankton (<0.22 m, <3 m, <8 m, <10 m, <40 m, <64 m, <112 m and <200 m) were estimated in 6 ponds and 5 experimental enclosures. The results showed that the planktonic algae less than 10 m are important in the biomass and production of phytoplankton in saline–alkaline ponds. The production of size fractionated phytoplankton corresponding to <112 m, <10 m and <3 m in saline–alkaline ponds were 10.5 ± 6.6 , 8.6 ± 5.4 and 0.33 ± 0.1 mgC l^–1 d^–1, respectively. Mean community respiration rate was 1.80 ± 0.73, 1.69 ± 0.90 and 1.38 ± 1.12 mgC l^–1 d^–1, respectively. The average production of phytoplankton corresponding to micro- (10–112 m), nano- (3–10 m) and pico- (<3 m) were 1.61, 8.30 and 0.33 mgC l^–1 d^–1, respectively. The ratio of those to the total phytoplankton production was 15%, 79% and 3%, respectively. The mean respiration rate of the different size groups was 0.11, 0.31 and 1.38 mgC l^–1 d^–1; the ratio of those to total respiration of phytoplankton was 6%, 17% and 77%, respectively. The production of size-fractionated phytoplankton corresponding to <200 m, <10 m and <3 m in enclosures was 2.19 ± 1.63, 2.08 ± 1.75 and 0.22 ± 0.08 mgC l^–1 d^-1, respectively. Mean community respiration rates were 1.25 ± 1.55, 1.17 ± 1.42 and 0.47 ± 0.32 mgC l^–1 d^–1, respectively. The average production of phytoplankton corresponding to micro- (10–200 m), nano- (3–10 m) and pico- (<3 m) plankton was 0.11, 1.86 and 0.22 mgC l^–1 d^–1, respectively. The ratio of those to the total production of phytoplankton was 5%, 85% and 10%, respectively. The mean respiration rate of different size groups were 0.08, 0.72 and 0.46 mgC l^–1 d^–1, the ratio of those to total respiration of phytoplankton was 6%, 57% and 37%, respectively. The concentrations of chlorophyll-a of the phytoplankton in the corresponding size of micro- (10–112 m), nano- (3–10 m) and pico- (<3 m) plankton in the experimental ponds were 19.3, 98.2 and 11. 9 g l^–1, respectively. The ratio of those to the total chlorophyll-a was 15%, 76% and 9%, respectively. The concentrations of chlorophyll-a of phytoplankton micro- (10–200 m), nano- (3–10 m) and pico- (<3 m) plankton in enclosures were 1.7, 34.3 and 3.0 g l^–1, respectively. The ratio of those to the total chlorophyll-a was 4%, 88% and 8%, respectively.     

Purification and characterization of two intracellular β-glucosidases from the Neurospora crassa mutant cell-1

Two intracellular -glucosidases (E.C. 3.2.1.21) were purified from the filamentous fungus Neurospora crassa, mutant cell-1 (FGSC no. 4335) and characterized. The extent of purification were 2.55- and 28.89-fold for -glucosidase A and -glucosidase B, respectively. -Glucosidase A was a dimeric protein, and B a monomeric protein, with molecular masses of 178 and 106 kDa, respectively. Both isoenzymes were glycoproteins with relatively high carbohydrate contents (-glucosidase A, 29.2%; -glucosidase B, 34.2%). The isoelectric points determined by IEF were 6.27 and 4.72, respectively. pH optima for activity were determined to be 5.0 and 5.5, and temperature optima to be 55 and 60 °C, for -glucosidases A and B, respectively. Both purified -glucosidases. especially -glucosidase B, showed relatively high stability against pH and temperature. Both enzymes were stable in the pH range of 5.0–9.0. The activities were completely retained up to 48 h at temperatures below 40 °C. At higher temperatures, enzymes were relatively unstable and lost their activities at 60 °C after 24 h. Both -glucosidases were highly activated by CuCl₂, and inhibited by SnCl₂ and KMnO₄. Hg²⁺ and Ag⁺ also inhibited severely -glucosidase B. The K _m and V _max values of the isoenzymes against cellobiose as substrate were 1.50 mM and 12.2mol min^–1 mg^–1 for -glucosidase A and 2.76 mM and 143.5 mol min^–1 mg^–1 for -glucosidase B.     

Detection and partial characterization of two antigenically distinct β-amylases in developing kernels of wheat

A -amylase (EC 3.2.1.2) was identified in the outer pericarp (P) of developing seeds of wheat (Triticum aestivum L.) and compared with the well known -amylase which is synthesized during seed development in the starchy endosperm (E). The enzyme P already exists in the tissues before anthesis and vanishes at the time when E starts to accumulate. The isoelectric-focusing patterns of P and E are very similar. The relative molecular weight (M_r) of P is slightly higher than that of E (66 and 64.5 kDa, respectively). Both P and E exhibit common epitopes in addition to epitopes specific for each of them. The two enzymes were identified in small amounts in the green tissues of the developing seeds (inner pericarp and testa). No antigenic difference was detected between P and the -amylases of roots and leaves.Abbreviations P pericarp -amylase - E endosperm -amylase - IS1 anti--amylase immune serum - IS2 anti- and anti- amylase immune serum - IS3 anti- amylase immune serum - IEF isoelectric focusing - IgG immunoglobulin G The authors thank Dr. P. Ziegler (Universität Bayreuth, FRG) for stimulating discussion and for useful suggestions during the writing of the text. The authors thank Miss C. Mayer for her skillful technical assistance.     

Exploration and exploitation of soil by apple,kiwifruit, peach,Asian pear and grape roots

Measurements of root-length density (RLD) in a range of 31 apple, kiwifruit, peach, Asian pear and grape orchards were used to derive indices to describe the exploration and exploitation of rooting volumes. Orchards were of various ages and located on a range of soil types, geographic regions, management systems etc. Data were obtained from core samples of volume 1.66×10^-4 m³ randomly taken within a standard volume, determined by average planting grids, of 2 m radius centred on tree stems, and 1 m depth. Root systems were described using an exploitation index, E(), and an exploration index, E(0). E() is defined as the proportion of the soil volume which contains roots at RLD greater than or equal to some specified value, . E(0) is defined as the proportion of the soil volume which contains roots at any RLD greater than zero. These indices are dependent on sample size, as are all volumetric or soil-coring data.Estimates of E(0) for each orchard were obtained as the proportions of cores containing any RLD>O and assessed for dependence on species. Peach trees had a significantly higher value of E(0), equal to almost 1.0, compared to the other four species where E(0) was approximately 0.8 (p0.01) or less. There was also some variation with age. E(0) was lower for very young plants which had not fully occupied the sampled soil volume. Exploration indices for woody roots increased with rootstock age but otherwise did not explain large differences in E() between species for given values.For example at =0.05×10⁴ m.m^-3, E() was approximately 0.45 for peach and kiwifruit, and 0.05 for apple, Asian pear and grape, whereas at =0.5×10⁴ m.m^-3 the corresponding values were 0.1 and almost zero. Negative exponential curves relating E(), scaled by dividing by E(0), to were fitted for each of the 31 orchards. Exponents for these curves, k, were significantly smaller for kiwifruit and peaches than apples, grapes and Asian pears (p0.05), and smaller for apples than grapes and Asian pears (p0.05). A larger k implies a rapid fall-off in E() as increases. Although all five species contained zero and low RLD samples, only kiwifruit and peaches contained higher RLD values and consequently have higher mean RLD. This trend was consistent across all soils, regions, sampling dates, and plant ages.The analyses demonstrate that core sampling can give useful insights into macro-scale root-system distribution, such as the proportion of a soil volume explored and how it is exploited. If positions of core samples are noted during sampling using angular direction, depth and radial distance as spatial coordinates the method can be used to describe root-system structures.     

Regulation of exocytosis in electrically permeabilized insulin-secreting cells. Evidence for Ca2+ dependent and independent secretion

The regulation of insulin secretion from RINm5F cells exposed to high voltage discharge has been investigated. Electron microscopy revealed that the overall structure of the cells was preserved after permeabilization. In this preparation insulin release was stimulated by Ca²⁺ (EC₅₀=2.4 M). The stable GTP analogue GTPS enhanced secretion both at intermediate (nano- to micromolar) and vanishingly low (<10 pM) Ca²⁺ concentrations. At optimal Ca²⁺ (10 M) the effect of GTPS was greatly reduced. We investigated whether the secretory response to GTP analogues was mediated by any of three enzyme systems regulated by GTP-binding proteins, i.e. generation of cyclic AMP by adenylate cyclase, of diacylglycerol by phospholipase C and of arachidonic acid by phospholipase A₂. The involvement of these messenger systems could be excluded as (i) cyclic AMP only had minor, Ca²⁺ dependent effects, (ii) phospholipase C was not activated in the absence of Ca²⁺ and insulin secretion due to the phorbol ester TPA displayed a different Ca²⁺ dependency, (iii) arachidonic acid did not elicit Ca²⁺ independent insulin secretion. These results, taken together with the finding that insulin secretion due to Ca²⁺ or TPA is attenuated by the inhibitory guanine nucleotide GDPS, suggest the existence of a regulatory site in exocytosis which is sensitive to guanine nucleotides.Abbreviations InsP₃ inositol trisphosphate - Ptd-InsP₂ phosphatidylinositol 4,5-bisphosphate - GTPS guanosine 5-(3-O-thio)triphosphate - GDPS guanosine 5-(2-O-thio)diphosphate - Gpp(NH)p guanyl-5-yl imidodiphosphate - TPA 12-O-tetradecanoylphorbol-13-acetate - OAG 1-oleoyl-2-acetylglycerol - Hepes 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid - EGTA (ethylenebis(oxyethylenenitrilo)tetraacetic acid - DAG diacylglycerol - [Ca²⁺]_i cytosolic free Ca²⁺ concentration     

Isolation and characterization of β-glucosidases from Aspergillus nidulans mutant USDB 1183

Two extracellular -glucosidases (cellobiase, EC 3.2.1.21), I and II, from Aspergillus nidulans USDB 1183 were purified to homogeneity with molecular weights of 240,000 and 78,000, respectively. Both hydrolysed laminaribiose, -gentiobiose, cellobiose, p-nitrophenyl--L-glucoside, phenyl--L-glucoside, o-nitrophenyl--L-glucoside, salicin and methyl--L-glucoside but not -linked disaccharides. Both were competitively inhibited by glucose and non-competitively (mixed) inhibited by glucono-1,5-lactone. -Glucosidase I was more susceptible to inhibition by Ag⁺ and less inhibited by Fe²⁺ and Fe³⁺ than -glucosidase II.     

Activation of peritoneal macrophages in patients with gynecological malignancies by sizofiran and recombinant interferon-γ

We investigated the influence of the combined use of sizofiran, a-1,3-glucan and a recombinant interferon- (rIFN-) upon biological activities of peritoneal macrophages (M). The number of peritoneal M and the production of cytokines (interleukin-1, interferon- and tumor necrosis factor) was increased by the combined treatment. Fully activated peritoneal M based on the increased number of elongated pseudopods were observed by electromicroscope. Sizofiran seems to assure a sufficient supply of M to kill tumor cells in the peritoneal cavity and co-administered rIFN- seems to directly stimulate the accumulated M in addition to its direct cytotoxicity against tumor cells. This combination therapy may be a step to the prevention of the recurrence of gynecological malignancies including ovarian cancer, after a negative second-look laparotomy.Abbreviations rIFN- recombinant interferon- - IL-1 interleukin-1 - TNF tumor necrosis factor - SLL second look laparotomy     

Characterization of Four Novel Variants of Goat βA-Globin Gene

Four novel alleles of the adult -globin gene of Capra hircus were observed in an extended study on hemoglobin polymorphism in goat breeds living in the island of Sardinia. Nucleotide sequencing showed that one of these alleles is due to a 2 bp substitution at codon 125 ( G, "LeuGlu). Two substitutions, the silent CT for Leu at codon 78 and the conservative A G (Lys Arg) at codon 104, are shared by the other three alleles, two of them having additional mutations, which suggests a common origin. The allele we provisionally called the ^Y shares four out of five amino acid substitutions, together with the same polymorphisms in the IVSII, we observed previously in the rather common ^E gene. This evidence allowed the origin of the ^E gene to be better characterized. The data increase to seven the number of alleles at the goat ^A -globin locus characterized thus far at the molecular level. A simplified nomenclature for the increasing number of goat -globin alleles is presented.     

First investigation of ultraoligotrophic alpine Lake Puma Yumco in the pre-Himalayas,China

Lake Puma Yumco is a typical alpine lake (altitude 5030m) located in the pre-Himalayas of Tibet, China, and this study was the first limnological investigation ever conducted on it. Lake Puma Yumco (28°34N, 90°24E) has the following morphometric properties: maximum length 31km, maximum width 14km, mean width 9km, shoreline 90km, surface area 280km², and shoreline development 1.5. Transparency was approximately 10m, even in the thawing season. The extinction coefficient of the lake water was calculated as 0.15m^–1. Annual maximum transparency was estimated from the depth of the Chara zone to be 30m. Dissolved oxygen was 7mg O₂ l^–1 and showed saturated values, and salinity was 360mgl^–1. The chemical type of the lake water was Mg-Ca-HCO₃-SO₄, and it was slightly alkaline in character. Total nitrogenous nutrients (sum of ammonia, nitrite, nitrate, and urea nitrogen), phosphate, and silicate were extremely low at 1, 0.02, and 9µM, respectively. Dissolved organic carbon, nitrogen, and phosphorus concentrations were 160, 11, and 0.08µM and the molar ratio was calculated as 2100:140:1. Chlorophyll a concentration was 0.2mgm^–3. Phytoplankton and zooplankton were dominated by Aphanocapsa sp. and Diaptomidae. Both nitrogen and phosphorus appear to be the limiting parameters for phytoplankton growth. Organic carbon and nitrogen contents in lake sediments were low and the sediments contained a large amount of CaCO₃. The grain size of sediment was that of silt-sand in most cases. The present results indicate that the pre-Himalayan alpine freshwater Lake Puma Yumco is an ultraoligotrophic lake.     

Ca2+-antagonists inhibit the N-methyltransferase‐dependent synthesis of phosphatidylcholine in the heart

Evidence indicates that, in addition to the Ltype Ca²⁺ channel blockade, Ca²⁺antagonists target other functions including the Ca²⁺pumps. This study was conducted to test the possibility that the reported inhibition of heart sarcolemmal (SL) and sarcoplasmic reticular (SR) Ca²⁺pumps by verapamil and diltiazem could be due to druginduced depression of phosphatidylethanolamine (PE) Nmethylation which modulates these Ca²⁺transport systems. Three catalytic sites individually responsible for the synthesis of PE monomethyl (site I), dimethyl (site II) and trimethyl (phosphatidylcholine (PC), site III) derivates were examined in SL and SR membranes by employing different concentrations of SadenosylLmethionine (AdoMet). Total methyl group incorporation into SL PE, in vitro, was significantly depressed by 10^–6–10^–3 M verapamil or diltiazem at site III. The catalytic activity of site I was inhibited by 10^–3 M verapamil only, whereas the site II activity was not affected by these drugs. The inhibition induced by verapamil or diltiazem (10^–5 M) was associated with a depression of the V_max value without any change in the apparent affinity for AdoMet. Both drugs decreased the SR as well as mitochondrial PE Nmethylation at site III. A selective depression of site III activity was also observed in SL isolated from hearts of rats treated with verapamil in vivo. Furthermore, administration of [³H-methyl]methionine following the treatment of animals with verapamil, reduced the synthesis of PC by Nmethyltransferase. Verapamil also depressed the N-methylation-dependent positive inotropic effect induced by methionine in the isolated Langendorff heart. Both agents depressed the SL Ca²⁺pump and although diltiazem also inhibited the SR Ca²⁺pump, verapamil exerted a stimulatory effect. In addition, verapamil decreased SR Ca²⁺-release. These results suggest that verapamil and diltiazem alter the cardiac PE Nmethyltransferase system. This action is apparently additional to the drugs' effect on Ltype Ca²⁺ channels and may serve as a biochemical mechanism for the drugs' inhibition of the cardiac Ca²⁺pumps and altered cardiac function.     

In vitro analysis of ion channels in periaxolemmal-myelin and white matter clathrin coated vesicles: Modulation by calcium and GTPγS

This study reports the analysis of K⁺ channel activity in bovine periaxolemmal-myelin and white matter-derived clathrin-coated vesicles. Channel activity was evaluated by the fusion of membrane vesicles with phospholipid bilayers formed across a patch-clamp pipette. In periaxolemmal myelin spontaneous K⁺ channels were observed with amplitudes of 25–30, 45–55, and 80–100 pS, all of which exhibited mean open-times of 1–2 msec. The open state probability of the 50 pS channel in periaxolemmal-myelin was increased by 6-methyldihydro-pyran-2-one. Periaxolemmal-myelin K⁺ channel activity was regulated by Ca²⁺. Little or no change in activity was observed when Ca²⁺ was added to thecis side of the bilayer. Addition of 10 M total Ca²⁺ also resulted in little change in K⁺ channel activity. However, at 80 M total Ca²⁺ all K⁺ channel activity was suppressed along with the activation of a 100 pS Cl^– channel. The K⁺ channel activity in periaxolemmal myelin was also regulated through a G-protein. Addition of GTPS to thetrans side of the bilayer resulted in a restriction of activity to the 45–50 pS channel which was present at all holding potentials. Endocytic coated vesicles, form in part through G-protein mediated events; white matter coated vesicles were analyzed for G proteins and for K⁺ channel activity. These vesicles, which previous studies had shown are derived from periaxolemmal domains, were found to be enriched in the subunits of G₀, G_s, and G_i and the low molecular weight G protein,ras. As with periaxolemmal-myelin treated with GTPS, the vesicle membrane exhibited only the 50 pS channel. The channel was active at all holding potentials and had open times of 1–6 msec. Addition of GTPS to the bilayer fused with vesicle membrane appeared to suppress this channel activity at low voltages yet induced a hyperactive state at holding potentials of 45 mV or greater. The vesicle 50 pS K⁺ channel was also activated by the 6-methyl-dihydropyron-2-one (20 M).Abbreviations CNPase 2–3 cyclic nucleotide phosphohydrolase - EDTA ethylenediamine N,N,N,N-tetraacetic acid - G-protein GTP(guanosine triphosphate) binding protein - GTPS guanosine 5-O-(3-thiotriphosphate) - MAG myelin associated glycoprotein - Na⁺ K⁺ ATPase, Na⁺ and K⁺ stimulated adenosine triphosphatase - PLP myelin proteolipid protein Special issue dedicated to Dr. Majorie B. Lees.     

A model for light adaptation: Producing Weber's law with bleaching-type kinetics

An adaptation model having two stages is introduced and its mathematical properties are examined. The two stages are the adaptive process (parameter K _b), which has bleaching-type kinetics, and the response function (parameters K _r and n), which incorporates response saturation. In order to study the increment threshold functions generated by the adaptation model the concept of a detector is required. It is demonstrated that without an adaptive process the compression hypothesis, in the form of the difference equation, produces increment threshold functions which saturate and do not obey Weber's law. It is then shown that an adaptive process with bleaching-type kinetics can prevent saturation and produce Weber's law behavior provided that the adaptive strength of the system exceeds the detector sensitivity.     

Some characteristics of anion transport at the tonoplast of oat roots,determined from the effects of anions on pyrophosphatedependent proton transport

The effects of anions on inorganicpyrophosphate-dependent H⁺-transport in isolated tonoplast vesicles from oat (Avena sativa L.) roots were determined. Both fluorescent and radioactive probes were used to measure formation of pH gradients and membrane potential in the vesicles. Pyrophosphate hydrolysis by the H⁺-translocating pyrophosphatase was unaffected by anions. Nonetheless, some anions (Cl^-, Br^- and NO₃-) stimulated H⁺-transport while others (malate, and iminodiacetate) did not. These differential effects were abolished when the membrane potential was clamped at zero mV using potassium and valinomycin. Stimulation of H⁺-transport by Cl^- showed saturation kinetics whereas that by NO₃- consisted of both a saturable component and a linear phase. For Cl^- and NO₃-, the saturable phase had a K _m of about 2 mol·m^-3. The anions that stimulated H⁺-transport also dissipated the membrane potential (.) generated by the pyrophosphatase. It is suggested that the stimulatory anions cross the tonoplast in response to the positive generated by the pyrophosphatase, causing dissipation of and stimulation of pH, as expected by the chemiosmotic hypothesis. The work is discussed in relation to recent studies of the effects of anions on ATP-dependent H⁺-transport at the tonoplast, and its relevance to anion accumulation in the vacuole in vivo is considered.Abbreviations and symools BTP 1,3-bis[tris(hydroxymethyl)-methylamino]-propane - EGTA ethylene glycol-bis(-aminoethyl ether)-N,N,N,N-tetraacetic acid - Hepes 4-(2-hydroxyethyl)-1-piperazine ethanesulphonic acid - IDA iminodiacetate - membrane potential - pH pH gradient - PPase inorganic pyrophosphatase - PPi morganic pyrophosphate     

Further studies on cell adhesion based on Lex-Lex interaction,with new approaches: embryoglycan aggregation of F9 teratocarcinoma cells,and adhesion of various tumour cells based on Lex expression

We previously proposed specific interaction of Le^x (Gal1 4[Fuc1 3]-GlcNAc1 3Gal) with Le^x as a basis of cell adhesion in pre-implantation embryos and in aggregation of F9 teratocarcinoma cells, based on several lines of evidence (Eggenset al., J Biol Chem (1989)264:9476–9484). We now present additional evidence for this concept, based on autoaggregation studies of plastic beads coated with glycosphingolipids (GSLs) bearing Le^x or other epitopes, and affinity chromatography on Le^x-columns of multivalent lactofucopentaose III (Le^x oligosaccharide) conjugated with lysyllysine. Comparative adhesion studies of Le^x-expressing tumour cellsvs their Le^x-non-expressing variants showed that only Le^x-expressing cells adhere to Le^x-coated plates and are involved in tumour cell aggregation, in analogy to F9 cell aggregation. The major carrier of Le^x determinant in F9 cells is not GSL but rather polylactosaminoglycan (embryoglycan), and we demonstrated autoaggregation of purified embryoglycan in the presence of Ca²⁺, and reversible dissociation in the absence of Ca²⁺ (addition of EDTA). Defucosylated embryoglycan did not show autoaggregation under the same conditions. Thus, Le^x-Le^x interaction has been demonstrated on a lactosaminoglycan basis as well as a GSL basis. A molecular model of Le^x-Le^x interaction based on minimum energy conformation with involvement of Ca²⁺ is presented.Abbreviations BSA bovine serum albumin - CHO carbohydrate - DMEM Dulbecco's modified Eagle's medium - EDTA ethylenediaminetetraacetic acid - GP glycopeptide - GSL glycosphingolipid - LAG lactosaminoglycan - Le^x Gal1 4[Fuc-1 3]GlcNAc1 R - LFP lacto-N-fucopentaose - LysLys-OH lysyllysinol - Mr relative molecular weight - PBS phosphate-buffered saline - PG paragloboside (Gal1 4GlcNAc1 3Gal1 4Glc1 1Cer) - TBS Tris-buffered saline (10mM Tris-HCl, pH 7.4, containing 0.15M NaCl) - TC tumour cell     

The Synthesis and Antiviral Activity of Glycyrrhizic Acid Conjugates with α-D-Glucosamine and Some Glycosylamines

Glycyrrhizic acid and its 30-methyl ester were conjugated with 2-amino-1,3,4,6-tetra-O-acetyl-2-deoxy--D-glucopyranose, 2,3,4,6-tetra-O-acetyl--D-glucopyranosyl amine, 2,3,4-tri-O-acetyl--L-arabinopyranosyl amine, 2-acetamido-2-deoxy--D-glucopyranosyl amine, and -D-galactopyranosyl amine using N,N-dicyclohexylcarbodiimide and its mixtures with N-hydroxybenzotriazole. Structures of the conjugates were confirmed by IR, UV, ¹H, and ¹³C NMR spectroscopy. The glycoconjugate with the residues of 2-acetamido-2-deoxy--D-glucopyranosyl amine in the carbohydrate part of its molecule exhibited antiviral activity (ID₅₀ 4 g/ml) toward the herpes simplex type 1 virus (HSV-1) in the VERO cell culture. Two compounds demonstrated anti-HIV-1 activity (50–70% inhibition of p24) in a culture of MT-4 cells at concentrations of 0.5–20 g/ml.