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1.
J F Collawn H Donato J K Upshur W W Fish 《Comparative biochemistry and physiology. B, Comparative biochemistry》1985,81(4):901-904
Ferritin was isolated from human liver and spleen. Reversed phase high performance liquid chromatography of the ferritin subunits from each tissue yielded the same three chromatographic fractions. Physical and chemical characterization of the three fractions indicated that they represented at least two, perhaps three, chemically distinct subunits. 相似文献
2.
James W. Drysdale 《The Biochemical journal》1974,141(3):627-632
1. Horse spleen ferritin and human liver ferritin were examined by gel electrofocusing under conditions that demonstrated equilibrium focusing. Both ferritins were resolved into multiple isoferritins. Both families of isoferritins were separable from one another. 2. Horse spleen ferritin was also resolved into five components by ion-exchange chromatography on DEAE-Sephadex A-50. Each of the major chromatographic fractions contained only a few of the isoferritins seen on gel electrofocusing. Each chromatographic fraction corresponded to different portions of the isoferritin profile. 3. These results indicate that the heterogeneity seen in many ferritins by gel electrofocusing represents structural heterogeneity in the ferritin population as isolated from the tissues. 相似文献
3.
J F Collawn L K Gowan H Crow C Schwabe W W Fish 《Archives of biochemistry and biophysics》1987,259(1):105-113
A partial amino acid sequence for three different subunits of the iron storage protein, ferritin, has been determined. Ferritin (Mr approximately 480,000) was isolated from porcine spleen and dissociated into its component subunits (Mr approximately 20,000). The subunits, in turn, were separated into three fractions by reversed-phase HPLC. The fractions appeared to be of equal size by sedimentation velocity, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and size-exclusion chromatography in 6 M guanidinium chloride. All three fractions were shown to be monomeric and to have no covalently attached carbohydrate (J. F. Collawn et al. (1984) Arch. Biochem. Biophys. 233, 260-266). Determination of the amino acid sequence of the C-terminal 70-80 residues from each of the fractions demonstrated three different sequences. Comparison with human liver H and L subunit sequences indicates that two of the porcine ferritin subunits are H-type subunits and one is an L-type subunit. Application of the Chou-Fasman algorithm on the three partial sequences suggests that these respective regions from each of the three subunits would probably adopt the same conformation. 相似文献
4.
Cell fractions were prepared from ACI rat livers and from rat hepatoma cell clone M-5123-C1. Radioimmunoassays of ferritin and of its protein subunits in various cell fractions after biosynthetic labeling with [14C]leucine were done by means of ferritin-specific and subunit-specific rabbit antibody. In both ACI rat livers and M-5123-C1 hepatoma cells free polyribosomes synthesized approximately 81% of the protein subunits of ferritin, and membrane-bound polyribosomes synthesized the rest. In both polyribosomal fractions, [14C]leucine-labeled subunits were detected earlier than [14C]leucine-labeled ferritin and apoferritin (5 min as against 30 min after initiation of a pulse). Time sequence studies of the shifts of biosynthetically labeled subunits and ferritin through different cell compartments provided evidence for vectorial transport of subunits and of ferritin, the direction of transport being from the two polyribosomal systems to the smooth membrane compartment and to the cytosol. 相似文献
5.
1. Human liver ferritin was separated by preparative isoelectric focusing into six fractions. 2. Except for the least acidic fraction the reactivity with antibody against spleen ferritin increased with rising pI, but with antibody against heart ferritin the reactivity decreased. 3. The highest iron content was found in the most acidic isoferritins and progressively decreased with rising pI. 4. Iron uptake was studied in apoferritin prepared from heart and liver ferritin fractions separated by ion-exchange chromatography. There was good correlation between the rate of iron uptake and pI. The most acidic fractions took up iron more rapidly than did the more basic ones. 5. Ferritin was prepared from heart, liver, spleen and kidney. There was little difference on isoelectric focusing between ferritin obtained from normal tissues and the corresponding iron-loaded tissues from patients who had received multiple blood transfusions. The iron-loaked heart ferritin invariably contained relatively more of the basic isoferritins. Normal and iron-overloaded heart ferritins were separated into isoferritin fractions by ion-exchange chromatography, and in each case there was a fall in iron content as the pI increased. The iron content of ferritin from the iron-overloaded heart was higher throughout than that from normal heart. 6. There is a relationship between the rate of iron uptake by apoferritin and pI, and this probably accounts for the variation in iron content of the isoferritins found in human liver and heart. 相似文献
6.
E J Frenkel B van den Beld J J Marx 《The International journal of biochemistry》1987,19(12):1229-1231
1. Purified rabbit liver ferritin was separated into fractions composed of 87% light molecular weight, and 72% high molecular weight subunits. 2. Radiolabeled ferritin fractions, injected intravenously into rabbits, showed no differences in clearance kinetics. 3. Differences found in ferritin clearance from plasma do not depend on the subunit composition of different molecular species. 相似文献
7.
Ferritin was purified from normal full-term placenta, and the native structure and subunit composition were characterized. Reversed-phase high-performance liquid chromatographic analysis of the placental ferritin subunits suggested the presence of three subunit types. Using acid urea gel electrophoresis and amino acid analysis, these subunits were tentatively identified as two H-type and one L-type. The relative proportions of the subunit types were approx. 23% H-1, 33% H-2 and 44% L. The native structure of placental ferritin as judged by circular dichroism and fluorescence spectroscopy was quite similar to that of ferritin isolated from horse spleen, a source that is composed predominantly of L subunits. These results are consistent with a ferritin tetracosameric structure whose H and L subunits fit into 24 equivalent sites interchangeably because the secondary and tertiary structures of the two subunit types are very similar. 相似文献
8.
Ascorbic acid inhibits lysosomal autophagy of ferritin 总被引:2,自引:0,他引:2
K R Bridges 《The Journal of biological chemistry》1987,262(30):14773-14778
Ascorbic acid retards ferritin degradation in K562 erythroleukemia cells leading to an increase in the availability of cellular iron (Bridges, K. R., and Hoffman, K. E. (1986) J. Biol. Chem. 261, 14273-14277). To explore the mechanism of this effect, the influence of ascorbate on subcellular ferritin distribution was examined. Cellular ferritin was pulse-labeled with 59Fe for 2 h, after which the cells were hypotonically lysed and fractionated on an 8% Percoll density gradient. Immediately after the labeling, all of the ferritin was in the cytoplasmic fractions at the top of the gradient. When the labeling was followed by a 24-h period of growth, a portion of the ferritin shifted to the lysosome-associated fractions at the bottom of the gradient, consistent with lysosomal autophagy of cytoplasmic ferritin. When ascorbate was added to the culture medium during the 24-h incubation, the magnitude of the shift was reduced. This process was also examined by size-fractionation of the contents of labeled cells using a Sepharose CL-6B column. Immediately after labeling, ferritin emerged from the column in two peaks, indicating the existence of both ferritin monomer and aggregates within the cytoplasm. After a 24-h period of growth, the monomer peak disappeared, while a new ferritin peak coincident with lysosomes emerged again, indicative of lysosomal autophagy of ferritin. In cells cultured with ascorbate for 24-h, there was a marked attenuation of the shift of ferritin to the lysosomal fractions. The monomer peak disappeared, as in the controls, but there was instead, an accumulation of ferritin as cytoplasmic aggregates. The total ferritin content of the ascorbate-treated cells was increased by 4-fold over that of the control. These experiments indicate that ascorbate blocks the degradation of cytoplasmic ferritin by reducing lysosomal autophagy of the protein. The access to the cell of the potentially toxic iron stored within the ferritin molecule is thereby increased. 相似文献
9.
Effect of ferritin-containing fractions with different iron loading on lipid peroxidation. 总被引:9,自引:5,他引:4
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J M Gutteridge B Halliwell A Treffry P M Harrison D Blake 《The Biochemical journal》1983,209(2):557-560
Ferritin-containing fractions with different degrees of iron loading were prepared. All ferritin fractions stimulated the peroxidation of bovine brain phospholipid liposomes, as measured by the formation of thiobarbituric acid-reactive material. This stimulation was increased in the presence of ascorbate. Iron salts of equivalent concentration to those of the ferritin fractions were more stimulatory to lipid peroxidation at the higher iron concentrations. None of the fractions inhibited ascorbate-dependent peroxidation in the presence of added iron salts. 相似文献
10.
S C Li H Kihara S Serizawa Y T Li A L Fluharty J S Mayes L J Shapiro 《The Journal of biological chemistry》1985,260(3):1867-1871
Urine specimens from two sibs affected with cerebroside sulfatase activator deficiency were examined to ascertain whether the deficiency of the supplementary activator protein required for the enzymatic hydrolysis of cerebroside sulfate was also evident in urine. Material from chromatographic fractionations was examined for the activator activity to avoid ambiguities resulting from protein inhibition. There were substantial deficits in all chromatographic fractions corresponding to activator-containing fractions of control urines. Since patient urines contained elevated amounts of lactosylceramide, digalactosylceramide, and globotriaosylceramide and since similarities between activators for cerebroside sulfate and GM1 ganglioside hydrolyses had been noted previously, the chromatographic fractions were also examined for activators in other glycosphingolipid hydrolase systems. There was coincidence of activators for the GM1 ganglioside/beta-galactosidase and the globotriaosylceramide/alpha-galactosidase A reactions with the cerebroside sulfatase activator in control urine fractions, and the patients' urines were deficient in activator activities for the three reactions. Identity of the three activators was suggested and antiserum to purified GM1 ganglioside activator was used to test this possibility. There were depressed levels of cross-reacting material in fractions of patient urines by Ouchterlony double diffusion and in unfractionated urine by enzyme-linked immunosorbent assay. Purified activators for the cerebroside sulfate and GM1 ganglioside systems showed lines of identity with no spurring on Ouchterlony double diffusion, identical mobility on immunoelectrophoresis, and similar stimulatory activities toward hydrolysis of the three glycosphingolipid species by their respective enzymes. Finally, the three activator activities were retained by anti-GM1-activator IgG coupled to Sepharose 4B. The results suggest strongly that the same protein entity serves as activator for the enzymatic hydrolysis of cerebroside sulfate, GM1 ganglioside, and globotriaosylceramide. 相似文献
11.
Development and optimization of a radioimmunoassay for duck serum ferritin described. 125I-labelled ferritin and rabbit anti-ferritin antibody are used together with goat anti-rabbit gamma globulin as separating agent for the bound and free fractions. The assay has a working range of up to 500 micrograms of ferritin per litre and a sample requirement of 50 microliter of serum. The assay requires a 24 h period and has a sensitivity of 10 micrograms of ferritin per litre. 相似文献
12.
Patricia L. Ringeling Maud I. Cleton Martin J. Kroos Lianne W. J. Sorber Wim C. de Bruyn Pauline M. Harrison Henk G. van Eijk 《Biometals》1989,2(2):114-121
Summary Cytosolic and lysosomal ferritin and haemosiderin were isolated from rat livers which had been iron-loaded by four intraperitoneal injections of iron-dextran. The cytosolic and lysosomal ferritins, prepared in a phosphate-free medium, were subjected to gel-filtration chromatography on Sepharose 613, yielding four fractions: a cytosolic monomeric (CMF) and void-volume ferritin fraction (CVVF), and a lysosomal monomeric (LMF) and void-volume ferritin fraction (LVVF). Of each fraction the following aspects were examined: (a) immunoreactivity against specific antiserum; (b) the Fe/P mass ratio and the effect of dialysis on this ratio using electron probe micro-analysis (EPMA); (c) morphology and Fespecific imaging using electron spectroscopic imaging (ESI) and electron energy loss spectroscopy (EELS). For haemosiderin one aspect, the Fe/P ratio, was determined before and after extensive purification. The following results were obtained (a) All ferritin fractions reacted with anti- (rat liver ferritin). (b) The Fe/P ratios as determined in CMF in an haemosiderin were not affected by dialysis or extensive purification, respectively. The Fe/P ratio in CWF was affected by dialysis. In the lysosomal fractions, only a trace of phosphorus (LVVF) or no phosphorus (LMF) was detected. (c) Morphologically, CMF and CVVF were found to be rather homogeneous; the iron core diameters of both fractions were in the known size range. LMF and LVVF were of rather heterogeneous composition; the core diameters of these fractions were different. In conclusion: the phosphorus in ferritin and haemosiderin is firmly bound; Haemosiderin, when derived from ferritin, has to take up phosphorus in the lysosomes. 相似文献
13.
Immuno-electron-microscopic studies on the subcellular distribution of rat liver epoxide hydrolase and the effect of phenobarbitone and 2-acetamidofluorene treatment. 总被引:1,自引:0,他引:1
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The distribution of rat liver epoxide hydrolase in various subcellular fractions was investigated by immuno-electron-microscopy. Ferritin-linked monospecific anti-(epoxide hydrolase) immunoglobulins bound specifically to the cytoplasmic surfaces of total microsomal preparations and smooth and rough microsomal fractions as well as the nuclear envelope. Specific binding was not observed when the ferritin conjugates were incubated with peroxisomes, lysosomes and mitochondria. The average specific ferritin load of the individual subcellular fractions correlated well with the measured epoxide hydrolase activities. This correlation was observed with fractions prepared from control, phenobarbitone-treated and 2-acetamidofluorene-treated rats. 相似文献
14.
Six peripheral blood lymphoid fractions (total lymphocytes, non-T, T, Tar (autologous rosette-forming T cells/precursor), T mu (helper), and T gamma (suppressor) lymphocytes) isolated through rosetting procedures were examined for the presence of ferritin by a direct immunofluorescence technique. Although ferritin was present in all lymphoid fractions studied, a significantly higher proportion of ferritin-containing cells were detected in the T-cell fraction than in the non-T-cell fraction, (mean +/- SD = 7.9 +/- 1.6% and 5.0 +/- 1.2%, respectively). T mu- and T gamma-cell fractions showed a twofold increase in the number of ferritin-positive cells (14.1 +/- 1.4% and 15.4 +/- 2.6%, respectively), as compared with Tar (7.0 +/- 0.9%)-and total lymphocyte (6.9 +/- 1.3%)-cell fractions. These results indicate that ferritin is preferentially distributed in T mu and T gamma lymphocytes and may constitute the basis for explaining some of the roles exercised by these cells in the control of other biological systems. 相似文献
15.
Ferritin and its protein subunits in rat hepatoma cell clone M-5123-C1 were biosynthetically labeled with [14C]leucine and 59Fe. Radioimmunoassays of ferritin/apoferritin and of protein subunits in the free polyribosome, membrane-bound polyribosome, smooth membrane, and cytosol fractions were done with ferritin-specific and subunit-specific rabbit IgG antibodies at various time intervals after pulsing. Much more 59Fe was bound by ferritin/apoferritin than by subunits in all of the cell fractions. Binding of iron to subunits may have been a random process. When hepatoma cells were simultaneously pulse-labeled with 59Fe and [14C]leucine, uptake of much of the 59Fe by ferritin occurred relatively early, in comparison to incorporation of [14C]leucine, in all of the cell fractions examined. Thus, 59Fe was readily incorporated into pre-existing ferritin. We conclude that most, if not nearly all, of the iron is incorporated after assembly of protein subunits. 相似文献
16.
Effects of starvation and different culture conditions on the phospholipid content of isolated pancreatic islets 总被引:3,自引:0,他引:3
A Hallberg 《Biochimica et biophysica acta》1984,796(3):328-335
Three forms of the normal human plasma fibrinogen gamma-chain which differ in molecular weight have been purified. Plasma fibrinogen was separated by ion exchange chromatography on DEAE-Sephacel into three populations of molecules, each with a unique gamma-chain composition. Following reduction and S-carboxymethylation, the fibrinogen polypeptide chains in each chromatographic peak were separated by ion exchange chromatography on DEAE-Sephacel and identified following sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The A alpha, B beta and smallest gamma-chain (gamma 50) eluted at progressively higher ionic strengths, but the elution positions of A alpha, B beta and gamma 50 chains were identical for fibrinogen from each of the three different chromatographic fractions. The unique gamma chain of fibrinogen in the second chromatographic peak (gamma 55) eluted at an ionic strength higher than that of the gamma 50 chain, while the largest gamma-chain (gamma 57.5), which was contained only in the third chromatographic peak of fibrinogen, eluted at the highest ionic strength. The higher ionic strengths needed to elute fibrinogen in the second and third peaks was paralleled by the higher ionic strengths needed to elute the gamma-chains unique to them, suggesting that the gamma-chain composition of the three fibrinogen fractions accounted for their differential binding to the ion exchange resin. Following desialation with neuraminidase, the differences in electrophoretic mobilities between the three gamma-chain forms was maintained, indicating that differential migration on SDS-polyacrylamide gel electrophoresis was not due to variation in sialic acid content. 相似文献
17.
Ferritin has been purified from normal full-term human placentae and its antigenic and molecular characteristics compared with adult liver ferritin. Placental ferritin is composed predominantly of a single subunit type, co-migrating with a liver ferritin standard on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. Comparison of dose-response curves in an immunoradiometric assay indicated some tissue-specific antigenicity for placental ferritin. This was supported by immunofluorescence studies on cryostat sections of human placentae by using antibodies to placental and spleen ferritin. Specific staining for placental ferritin was demonstrated within placental syncytiotrophoblast, particularly localized towards the microvillus plasma membrane. Ferritin has also been shown by electrophoretic and antigenic analysis to be present in protein fractions solubilized from isolated human syncytiotrophoblast microvillus plasma-membrane preparations, suggesting that ferritin may play an active role in the transfer of iron from maternal transferrin across the syncytiotrophoblast plasma membrane. 相似文献
18.
Patrick L. Storring Siddiqua Fatih 《Biochimica et Biophysica Acta (BBA)/General Subjects》1975,392(1):26-38
This paper describes a study of the incorporation of 5 9Fe from 5 9Fe-labelled rat transferrin into rat bone marrow cells in culture. 5 9Fe was found in both stroma and cytoplasm of marrow cells, and the cytoplasmic 5 9Fe separated by polyacrylamide gel electrophoresis, into ferritin, haemoglobin and a low molecular weight fraction.The incorporation of 5 9Fe into all three cytoplasmic fractions, but not into the stroma, increased progressively with time. Erythropoietin stimulated the increase of 5 9Fe in ferritin within 1 h, the earliest time examined, and more than 3 h later in the stroma and haemoglobin.A proportion of the 59Fe incorporated into the stroma and low molecular weight iron fractions during a 1 h incubation with 59Fe-labelled transferrin was mobilised into ferritin and haemoglobin during a subsequent 4-h “cold-chase”. Erythropoietin, when present during the “cold-chase”, did not influence these 59Fe fluxes. The erythropoietin stimulation of 59Fe incorporation into ferritin, one of the earliest erythropoietin effects to be recorded, was therefore considered to be due to an increase of 59Fe uptake by the hormone-responsive cells rather than a direct effect on ferritin synthesis.20-h cultures containing erythropoietin when incubated with 59Fe-labelled transferrin for 4 h, showed dose-related erythropoietin stimulation of 59Fe incorporation into haemoglobin only.In the presence of 10 mM isonicotinic acid hydrazide, 59Fe incorporation into haemoglobin was inhibited, as in reticulocytes (Ponka, P. and Neuwirt, J. (1969) Blood 33, 690–707), while that into the stroma, ferritin and low molecular weight iron fractions, was stimulated; there were no reproducible effects of erythropoietin. 相似文献
19.
The isolation and purification of fractions with juvenile hormone activity from whole animal extracts of larvae, and from extracts of haemolymph from larvae and adults is described. Using the Galleria bioassay three such fractions were isolated from both third and fourth instar hoppers and from adult females. The chromatographic behaviour of these fractions indicates that one contains JH III, but the other two contain unknown juvenilizing compounds. 相似文献
20.
Gel filtration and chromatographic separation of soil extracts gave three fractions which induced formation of chlamydospores by Fusarium solani f. sp. phaseoli. Depletion of nutrients had a similar effect. 相似文献