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1.
  • 1.1. Platelets bind specifically to lactoferrin. A significant similarity between human lactoferrin and some bovine milk proteins has been established.
  • 2.2. Because of the structural homology of lactoferrin and cows milk proteins they are able to influence lactoferrins regulatory function on the level of its binding to membrane receptors on platelets.
  • 3.3. An inhibitory effect of bovine α-lactalbumin and of β-lactoglobulin on lactoferrin-receptor interaction was shown.
  • 4.4. Bovine α-lactalbumin competes with lactoferrin for the binding sites.
  • 5.5. Scatchard plot analysis of data shows one binding site for lactoferrin in the presence of α-lactalbumin with an affinity constant, Ka = 0.46 × 109 mol/1 and 335 receptors/cell.
  • 6.6. The inhibitory effect of β-lactoglobulin reaches 62% and is different for the common fraction ⨿-lactoglobulin and the genetic variants β-lactoglobulin A and B.
  • 7.7. β-lactoglobulin does not compete with lactoferrin for the membrane receptors.
  • 8.8. Bovine casein and egg lysozyme stimulate 59Fe-lactoferrin binding to the receptors. The mechanism of these effects is still unknown.
  • 9.9. Tested alimentary antigens are able to interact with lactoferrin and also with some platelet membrane structures.
  • 10.10. Established changes in lactoferrin binding to the platelet membrane might be in relation to lactoferrins regulatory function and (or) eliminating mechanisms of these alimentary antigens.
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2.
  • 1.1. A non-radioisotopic method utilizing a biotin-avidin approach was used to characterize lactoferrin binding to the clonal MAC-T bovine mammary epithelial cell line.
  • 2.2. Binding of lactoferrin to MAC-T cells and isolated membranes was specific and saturable.
  • 3.3. Unlabeled lactoferrin competed for and displaced biotin-labeled lactoferrin from binding sites on mammary epithelial cells. In contrast, unlabeled transferrin did not compete.
  • 4.4. Scatchard analysis of lactoferrin binding to MAC-T cell crude membranes was nonlinear, revealing two classes of binding sites with association constants (Ka) of 2.36 × 107 and 3.36 × 106M−1.
  • 5.5. Binding of lactoferrin to MAC-T cells may be associated with the initial events which result in decreased MAC-T cell proliferation.
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3.
  • 1.1. A half platelet preparation from Chinese crab (Eriocheir sinensis) gill is described which allows electrophysiological investigations of ion transport by gill epithelial monolayer when mounted in a modified Ussing chamber.
  • 2.2. The resistance of these preparations equals half that of complete gill platelets (containing the gill epithelium and cuticle twice) indicating that cell damage during preparation of half platelets is negligible.
  • 3.3. The transepithelial resistance (resistance of cuticle subtracted previously) was determined to be about 140 Ω cm2 when both sides are bathed with identical salines.
  • 4.4. Similarities to the results obtained with perfused complete gills demonstrates the reliability of this preparation.
  • 5.5. When identical salines are applied on both sides of the epithelium an outside positive transepithelial potential difference (PDte) up to 40 mV was measured.
  • 6.6. The occurrence of such a high PDte under symmetric conditions and its sensitivity to CN suggests the PDte to be generated by active transport processes.
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4.
  • 1.1. The diffusional water permeability (Pd) of rabbit red blood cell (RBC) membrane has been monitored by a doping nuclear magnetic resonance (NMR) technique on control cells and following inhibition with p-chloromercuribenzene sulfonate (PCMBS).
  • 2.2. The values of Pd were around 6.3 × 10−3 cm/sec at 15°C, 7.0 × 10−3cm/sec at 20°C, 8.0 × 10−3 cm/sec at 25°C, 9.1 × 10−3 cm/sec at 30°C and10.7 × 10−3 cm/sec at 37°C.
  • 3.3. Systematic studies on the effects of PCMBS on water diffusion indicated that the maximal inhibition was reached in 15 min at 37°C with 0.5 mM PCMBS.
  • 4.4. The values of maximal inhibition were around 71–74% at all temperatures.
  • 5.5. The basal permeability to water was estimated as 1.6 × 10−3cm/sec at 15°C, 2.0 × 10−3cm/sec at 20°C, 2.4 × 10−3cm/sec at 25°C, 2.6 × 10−3cm/sec at 30°C, and 3.1× 10−3 cm/secat 37°C.
  • 6.6. The activation energy of water diffusion was around 18 kJ/mol and increased to 27 kcal/mol after incubation with PCMBS in conditions of maximal inhibition of water diffusion.
  • 7.7. The membrane polypeptide electrophoretic pattern of rabbit RBCs has been compared with its human counterpart.
  • 8.8. The rabbit membrane contained a higher amount of spectrin (bands 1 and 2), while the band 6 (glyceraldehyde-3-phosphate dehydrogenase) was markedly less intense.
  • 9.9. Considerable differences in the electrophoretic patterns of the two sources of RBC membranes appeared in the bands migrating in the band 4.5 region and in front of band 7, where some polypeptides were apparent in higher amounts in the rabbit RBC membrane.
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5.
  • 1.1. Insulin and insulin-like growth factor I (IGF-I) receptors were studied in bovine chromaffin cells isolated from the medulla by collagenase digestion and kept in primary culture.
  • 2.2. Specific 125I-labelled insulin binding increased with time in culture with no significant change in the dissociation constant, Kd~0.3nM. Insulin was nearly 100-fold more potent than IGF-I in displacing 125I-labelled insulin.
  • 3.3. Affinity crosslinking and SDS gel electrophoresis revealed increased binding of 125I-labelled insulin and 125I-IGF-I with time in culture, the densities of the labelling indicating relatively a much higher expression of IGF-I than insulin receptors in the cells. The apparent molecular weight of both the hormone binding subunits were 135,000, suggesting that the insulin and IGF-I receptors in the adrenal medulla are of the peripheral types.
  • 4.4. Both receptors thus appeared to be affected by the collagenase treatment but with a subsequent recovery when cells were kept in culture.
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6.
  • 1.1. The mitochondrial dihydropyridine receptor was solubilized with Chaps at a detergent/ protein ratio of 2.5, during 45 min at 4°C.
  • 2.2. From the rate constants of association (8.10 ± 0.25 × 104 M−1 min−1) and dissociation (0.022 ± 0.001 min−1 a Kd of 275 nM was calculated, while from saturation experiments a Kd of 270 ± 30 nM and a density of receptors of 106 ± 9 pmol/mg protein was obtained.
  • 3.4. The solubilized receptors are heat-resistant, sensitive to the trypsin and to the reduction of disulfide bonds.
  • 4.5. In native membranes, a polypeptide of 50 kDa was specifically photolabelled with [3H]Azidopine.
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7.
  • 1.1. We report for the first time on the production and characterization of antibodies against a naturally occurring tetrahydroisoquinoline, namely salsolidine (6,7-dimethoxy-1-methyl-1,2,3,4-tetrahydroisoquinoline).
  • 2.2. Immunogen synthesis was carried out by coupling the hapten salsolidine to bovine serum albumin (BSA) as carrier protein on the basis of reductive amination.
  • 3.3. By immunization of rabbits with salsolidine-BSA conjugate antisalsolidine antibodies were produced.
  • 4.4. At a final dilution of 1:1700 the highest-litre antiserum bound 35% of 0.21 pmol [3H] salsolidine. This antiserum was used to develop a radioimmunoassay for salsolidine.
  • 5.5. Cross-reactivity studies revealed a high specificity of the antiserum to the hapten.
  • 6.6. The antibodies had a high affinity to salsolidine (Ka = 1.5 × 109 M−1).
  • 7.7. Standard curves covered a measuring range of 0.5–70 pmol/tube and the detection limit was found to be 0.27 pmol/tube.
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8.
  • 1.1. A method for purifying undischarged nematocysts from Hydra and other cnidarians is described.
  • 2.2. Isolated cysts (relative densities 1.22–1.24) evaginate their tubular content even after previous dehydration.
  • 3.3. The cyst wall is permeable to dyes of mol. wts up to 600,000.
  • 4.4. Approximately two-thirds of the cyst's dry wt are soluble proteins. Eighty per cent of them are of low mol. wt and highly anionic, presumably serving as binding sites for Ca2+ and Mg2+.
  • 5.5. The other 20% includes 30 different proteins amongst them toxins and enzymes (phospholipase and little proteases but no collagenase, chitinase or hyaluronidase).
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9.
  • 1.1. The interaction of haemopexin and albumin with TPPS4 was studied by measuring the absorption and fluorescence spectra. Haemopexin was found to have one strong TPPS4 binding center (Ka = 3 × 107M−1).
  • 2.2. Haem-haemopexin complex appears to have no specific binding site for TPPS4. Occupation of the specific binding center of haemopexin molecule by a haem abolishes TPPS4 binding.
  • 3.3. Albumin was found to possess one strong TPPS4 binding center (Ka = 3 × 106M−1) besides two or three weak binding sites (Ka = 2 × 105M−1).
  • 4.4. Haern-albumin complex possesses only one weak TPPS4 binding site (Ka = 7 × lO5M−1). These observations suggest identity of primary binding sites of TPPS4 and haem on albumin molecule.
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10.
  • 1.1. This study examined the effect of the monoamines dopamine and octopamine, as well as tyrosine on the oxygen affinity and cooperativity of oxygen binding by the hemocyanin of the marine gastropod Busycon canaliculatum. The effect of temperature on hemocyanin oxygen affinity was also examined.
  • 2.2. Freezing Busycon hemocyanin did not affect the binding of oxygen.
  • 3.3. Dopamine, octopamine and tyrosine had no significant effect on the oxygen affinity or cooperativity of oxygen binding by the hemocyanin of B. canaliculatum.
  • 4.4. It was concluded that Busycon hemocyanin either has no binding sites for the two monoamines or for tyrosine, or that binding of the molecules has no functional significance.
  • 5.5. Both temperature sensitivity and affinity of hemocyanin-oxygen binding were similar to values previously reported for hemocyanin of Busycon from other localities.
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11.
  • 1.1. Increases in membrane conductance (gm) were induced by GABA in distal bundles 32, 33 and 34 of extensor tibiae muscles of the locust (Schistocerca gregaria).
  • 2.2. Bath application of GABA (10−5−5 × 10−3 M) induced reductions in muscle fibre space constant (λ).
  • 3.3. GABA (5 × 10−3 M) induced additional membrane conductance of 2.21 ± 0.03 × 10−6 S/mm, 0.38 ± 0.03 × 10−6 S/mm and 0.29 ± 0.06 × 10−6 S/mm on muscle bundles 34, 33 and 32 respectively. The greater sensitivity of muscle fibres in bundle 34 to GABA is due at least in part to a larger number of GABA receptors on bundle 34 muscle fibres.
  • 4.4. The decrement of electrotonic potentials in the presence of GABA were measured over distances of both half fibre length and whole fibre length. Good agreement was obtained between changes in space constant produced by GABA using half fibre length and whole fibre length data.
  • 5.5. By taking into account changes in space constant induced by GABA it was possible to demonstrate that presynaptic GABA receptors were involved in the inhibition of slow excitatory postsynaptic potentials by GABA.
  • 6.6. “Slow” excitatory postsynaptic potentials recorded under current clamp were inhibited in a dose-dependent manner by GABA. This inhibition was not dependent on muscle-fibre GABA sensitivity and could not be completely accounted for by GABA-induced changes in the cable properties of the muscle fibres.
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12.
  • 1.1. Kinetic and physico-chemical studies on human placental microsomal fraction confirmed that the ATPase and ADPase activities detected in this fraction correspond to the enzyme ATP-diphosphohydrolase or apyrase (EC 3.6.1.5). These include substrate specificity, and coincident Mr and pI values of both ATPase-ADPase activities.
  • 2.2. This enzyme hydrolyses both the free unprotonated and cation-nucleotide complex, the catalytic efficiency for the latter being considerably higher.
  • 3.3. Microsomal apyrase is insensitive to ouabain and Ap5A. The highly purified enzyme was only inhibited by o-vanadate, DBS and slightly by DCCD.
  • 4.4. Apyrase seems to be a glycoprotein from its interaction with Concanavalin-A.
  • 5.5. Preliminary studies on the essential amino acid residues suggest the participation of Arg, Lys and His residues, and discard the requirement of −SH, COO, −OH, and probably also Tyr and Trp.
  • 6.6. Two kinetic modulatory proteins of apyrase were detected in placental tissue. An activating protein was found in the soluble fraction and an inhibitory protein was loosely bound to the membranes.
  • 7.7. The proposed in vivo function for apyrase is related to the inhibition of platelet aggregation due to its ADPase activity, which is supported by the direct effect on washed platelets and by its plasma membrane localization.
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13.
  • 1.1. S-d-Lactoylglutathione accumulates in human platelets activated by agonists. Among the tested inducers thrombin is the most active.
  • 2.2. The effect is dose and time-dependent. S-d-Lactoylglutathione, corresponding to depleted pool of reduced glutathione, can also be detected in platelets incubated with exogenous methylglyoxal.
  • 3.3. A further significant increase was observed in platelets stimulated with trombin in the presence of methylglyoxal.
  • 4.4. No change in glyoxalase activities upon platelet stimulation with thrombin was shown.
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14.
  • 1.1. The O2-binding characteristics of the blood of the euterrestrial amphipod (landhopper) Arcitalitrus dorrieni have been studied.
  • 2.2. The blood exhibited a low O2 affinity, with a p50 (at pH = 7.8) of 21.4 torr (10°C). Affinity decreased with an increase in temperature at constant pH (ΔH = − 79.4kJ/mol) but the Bohr factor (ΔlogP50/Δ pH = −0.67) was unaffected.
  • 3.3. The O2-carrying capacity of the blood was moderate (1.51 ml/100 ml)
  • 4.4. The results support the hypothesis that the blood of terrestrial amphipods is characterized by having a low affinity pigment.
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15.
  • 1.1. Subcellular distribution of (NA+, K+-ATPase and ouabain-insensitive ATPase (Mg2+-ATPase) are compared in branchial tissues of the euryhaline crab, Eriocheir sinensis, acclimated to fresh water.
  • 2.2. Both the anterior and posterior gills contain cAMP-dependent protein kinase and endogenous protein substrate for phosphorylation.
  • 3.3. Phosphorylation occurs in both “particulate” and “soluble” subcellular fractions but its stimulation by cAMP is restricted to the “soluble” fraction.
  • 4.4. serotonin (5-HT) and dopamine receptors are present only in the “light particulate” fraction isolated from the posterior gills.
  • 1.(a) Serotonin and dopamine have no effect on the phosphorylation observed in a subcellular fraction alone.
  • 2.(b) Activation of the phosphorylation by serotonin and dopamine is found when the soluble fraction (source of cAMP-dependent protein kinase) is added to the fraction P3 from the posterior gills.
  • 3.(c) No activation occurs with the fractions P3 as well as P1 or P2 (not shown) from anterior gills of fresh water crab.
  • 4.(d) Cyproheptadine, a serotonin receptor antagonist, inhibits the 5-HT dependent increase in phosphorylation.
  • 5.(e) The dopamine receptor antagonist, chlorpromazine, inhibits dopamine-stimulated phosphorylation.
  • 6.5. Ouabain mimics the effect of cyproheptadine on the serotonin-stimulated phosphorylation found in the posterior gills.
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16.
  • 1.1. The binding of O2 to goldfish haemoglobin showed a strong pH dependence P50=5.5 mmHg; n = 2.4 at pH 8.0 and P50 = 170 mmHg; n = 1.0 at pH 5.5 such that the protein is only 50% saturated in a solution of air equilibrated buffer at pH 5.5.
  • 2.2. The binding of CO is cooperative at high pH (n = 2.8; L = 1000; KR = 0.1 μM; KT = 4 μM) and non-cooperative (n = 1) at pH 5.5.
  • 3.3. The rate of O2 dissociation is extremely fast and pH dependent; being 30 sec−1 at pH 8.0 and 400 sec−1 at pH 6.0 at 1°C. At 23°C the rate of this process is too fast to obtain accurate data using stopped-flow techniques.
  • 4.4. Partial photolysis of the oxyhaemoglobin species leads to homogeneous recombination kinetics at pH 8.0 with an associated rate constant of 4.7 × 107 M−1 sec−1. At pH < 7.5 the recombination process occurs in two steps. One rate is equal to that observed at pH 8.0. The slower process is favoured at low pH.
  • 5.5. Photolysis of the CO haemoglobin complex indicates that, at high pH, combination of CO with deoxyhaemoglobin is cooperative, whilst recombination with Hb(CO)3 is non-cooperative and occurs at a rate of 1.2 × 106 M−1 sec−1.
  • 6.6. At neutral pH recombination of CO with partially linganded haemoglobin occurs in a two-step process. The proportion contributed by each of these two steps in pH dependent.
  • 7.7. The functioning of this Root effect haemoglobin is discussed in terms of the two state-model of cooperativity in which the αβ chain heterogeneity is minimal
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17.
  • 1.1. The properties of Na+/K+-transporting ATPase in microsomal fractions from the nervous tissue of the grasshopper, Poekilocerus bufonius were investigated.
  • 2.2. Two components of ATPase activity are present.
  • 3.3. Inclusion of 1 mM ouabain in the incubation media reduced the activity of total and Na+/K+-ATPase by 57 and 79%, respectively.
  • 4.4. The maximum velocity (Vmax) was decreased by the addition of 1 mM ouabain, whereas the apparent Km value was not affected indicating a non-competitive type of inhibition.
  • 5.5. The calculated value of the pI50 was 6.4 (I50 = 3.98 × 10−7M) for ouabain inhibition of the enzyme showing great sensitivity to the cardiac glycoside ouabain.
  • 6.6. The present results show that the physicochemical properties of Na+/K+-transporting ATPase from the brain of P. bufonius are essentially the same as for the enzyme prepared from the excretory system of the insect which has been previously investigated.
  • 7.7. Dissimilarities were also observed between these tissues in the way that the enzyme from the brain was sensitive to ouabain inhibition with a non-competitive type rather than a ouabain-resistance and a competitive type of inhibition for the enzyme from the excretory system.
  • 8.8. These dissimilarities are probably due to different isoenzyme patterns available in the same insect.
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18.
  • 1.1. The enzyme fructose-1,6-bisphosphatase was purified from the mantle of the sea mussel Mytilus galloprovincialis Lmk. The purified enzyme showed a single band in SDS-polyacrylamide gel electrophoresis. The mol. wt and subunit mol. wt of the enzyme were 105,000 and 27,000, respectively.
  • 2.2. Divalent cations are essential for the enzyme activity. In the absence of chelating agents, FBPase 1 exhibits hyperbolic kinetics with respect to Mn2+, Zn2+ and Mg2+. The Km for Mg2+ is lower than the physiological concentration of cation in the tissue, whereas its Km for Mn2+ and Zn2+ is greater than the respective in vivo concentrations.
  • 3.3. The joint action of Mg2+ and Zn2+ increases the affinity of the enzyme for the substrate Fru-1,6-P2, though Vmax is reduced.
  • 4.4. Na+ strongly inhibits the enzyme even at very low concentrations. K+ has no effect whatsoever.
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19.
  • 1.1. The acid phosphatase (AcPase, EC 3.1.3.2) IV from rat testicular tissue was purified to apparent homogeneity.
  • 2.2. The enzyme displays a native molecular weight of 70 kDa determined on gel permeation chromatography on a Sephadex G-100 column and 68 kDa using linear 5–20% sucrose density gradient centrifugation. The subunit molecular weight on SDS-PAGE analysis is 67 kDa, suggesting that the enzyme is a monomeric protein.
  • 3.3. The enzyme does not bind to Concanavaline A-Sepharose 4B column, indicating that it is not a glycoprotein.
  • 4.4. The rat testis AcPase IV is a metal activated enzyme in which Mg2+ is the metal activating agent with a Ka, = 0.88 × 10−3 M. The Michaelis constant for p-nitrophenylphosphate, in the presence of saturating concentrations of Mg2+ ions, is 0.23 × 10−3 M.
  • 5.5. The enzyme preferentially hydrolizes p-nitrophenylphosphate, phenylphosphate and ATP.
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20.
  • 1.1. The copepod Acartia clausi exhibited two laminarinases (exo- and endo-acting forms) purified by gel chromatography followed by affinity chromatography. Specific antibodies have been raised against the purified exolaminarinase antigen.
  • 2.2. A single band of protein appeared on a polyacrylamide disc gel electrophoresis; its mol. wt is 21,000.
  • 3.3. Biochemical properties of the purified enzyme showed a maximum activity at pH 5.2 and a temperature of 40°C with laminarin as substrate. The thermal stability of the enzyme and the effect of various cations on its activity were examined. The enzyme hydrolyses specifically the β(1–3) linked polysaccharides and had no activity against the α(1–4) or β(1–4) disaccharides or polysaccharides.
  • 4.4. The kinetic parameters Vm and Km vary with the temperature; the affinity constant (Ka) was maximum between 25–30°C. The Arrhenius plot defined two values of energy of activation: 7980 cal/mole and 17,506 cal/mole.
  • 5.5. From the purification scheme the exoacting form appears to be largely dominant over the endoacting form.
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