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1.
M. Böttger L. Karawajew H. Fenske K. Grade R. Lindigkeit 《Molecular biology reports》1981,7(4):231-234
Salt-dependent structural changes of rat liver chromatin isolated by an extraction procedure not involving shear and exogeneous nucleases were investigated by sedimentation and light scattering methods. The effects observed are complex involving changes of the molecular weight and expansion. Between 0.1 M and 0.2 M (NH4)2SO4 where histone H1 is released, a fragmentation into molecules of half molecular weight is found which is accompanied by an expansion into a more extended conformation gradually increasing to 0.4 M (NH4)2SO4. The H1-free chromatin does not exhibit the reduction in molecular weight but undergoes this expansion. The original conformation is not reversible on re-decreasing the salt concentration to 0.1 M (NH4)2SO4. 相似文献
2.
Along with proteins, lipids, water and minerals, polysaccharides are the main chemical compounds of which macroalgae are built. Among the chemical compounds now widely examined is fucoidan (fucan, fucosan, sulfate fucan or sulfated fucan), a fucose-containing sulfated polysaccharide. Fucoidans isolated from different species have been extensively studied because of their varied biological properties, including anticoagulant and antitumor effects. Methodology based on mild acid hydrolysis can be used as an efficient tool to study the relationship between molecular weight of the sulfated polysaccharides and their biological activities. Anticancer activity of fucoidans can be significantly enhanced by lowering their molecular weight only when they are depolymerized under mild conditions. In this study, fucoidan was identified during extraction with H2SO4 and HCl; its presence was confirmed by FT-Raman spectroscopy in aqueous solution. In particular, shifts at 840 cm−1 were analysed, which are due to the presence of sulfate at the axial C-4 position, as were the shifts at about 811–809 cm−1, for which the sulfated fucoidan is responsible. Shifts of electrophoretic bands of fucoidan resulting from mild acid hydrolysis in H2SO4 and HCl were also analysed. The analytical procedure was developed using apparatus for cellulose acetate membrane electrophoresis and this was supplemented by semi-quantitative analysis. 相似文献
3.
Criteria for assessing the precision and accuracy of methods for estimation of molecular weight for proteins using sodium dodecyl sulfate-polyacrylamide-gel electrophoresis have been applied to rhodopsin from bovine visual cell outer segment membranes. Various methods of preparing this hydrophobic protein for electrophoresis differ in their ability to solubilize and disaggregate polypeptide constituents of the outer segment membrane, with resultant variations in the pattern of protein bands and the apparent molecular weight of rhodopsin. Even with optimal solubilization and disaggregation, the behavior of rhodopsin relative to a series of standard proteins is such that the apparent molecular weight decreases systematically from 40,400 to 34,500 as the acrylamide concentration increases from 4 to 10%. As demonstrated by Ferguson plots of logRf vs gel concentration and split gel experiments, this discrepancy is explained by the fact that the extrapolated Rf for zero gel concentration (Y0) for rhodopsin is significantly lower than the Y0's for the soluble proteins used as molecular weight standards. In such cases, a possibly more reliable molecular weight estimate is obtained by plotting the retardation coefficient (KR) vs molecular weight. This method yields a value of 29,500 ± 1000 for bovine rhodopsin if only the errors in measurement of Rf are considered and a quadratic relationship between KR and molecular weight is used. Using weighted linear regression for KR vs molecular weight, we obtain a molecular weight estimate of 32,700 ± 5000 when the uncertainty in the calibration curve is considered. Because of uncertainties regarding the detergent-binding properties of rhodopsin and the relationship of its Stokes radius to its molecular weight by comparison with the soluble protein standards, these values must be viewed with caution. 相似文献
4.
Flow dichroism of deoxyribonucleic acid solutions 总被引:1,自引:0,他引:1
The flow dichroism of dilute DNA solutions (A260 ≈ 0.1) has been studied in a Couette-type apparatus with the outer cylinder rotating and with the light path parallel to the cylinder axis. Shear gradients in the range of 5–160 sec.?1 were studied. The DNA samples were whole, “half,” and “quarter” molecules of T4 bacteriophage DNA, and linear and circular λb2b5c, DNA. For the linear molecules, the fractional flow dichroism is a linear function of molecular weight. The dichroism for linear λ DNA is about 1.8 that of the circular molecule. For a given DNA, the dichroism is an approximately linear function of shear gradient, but with a slight upward curvature at low values of G, and some trend toward saturation at larger values of G. The fractional dichroism increases as the supporting electrolyte concentration decreases. 相似文献
5.
M. Böttger H. Fenske L. Karawajew H. Hamann K. Karawajew R. Lindigkeit 《Molecular biology reports》1982,8(3):137-142
A chromatin fraction, which can reproducibly be extracted from rat liver nuclei at moderate salt concentration (0.1 M (NH4)2SO4, 0.1 M Tris-HCl, 2 mM MnCl2, pH 7.9), was analyzed with regard to changes of its molecular weight in the range of (NH4)2SO4 concentrations between 0.1 M and 0.4 M. With the transition from 0.1 M to 0.2 M (NH4)2SO4 histone H1 is released and the molecular weight obtained from both sedimentation-viscosity and light scattering is reduced by approximately one-half. A spatial expansion of the resulting half-molecules is observed with further increasing salt concentration. On the basis of these results a double-fibrillar structure of this chromatin fraction is proposed. 相似文献
6.
Hybrid magnetic Fe3O4@SiO2-poly(ethylene oxide)-maltose (Fe3O4@SiO2-PEO-mal) nanoparticles synthesized by our group can be used as affinity adsorption carriers for direct separation of maltose binding protein-fused Hep I (MBP-Hep I) from a crude enzyme solution in a magnetic field. In this work, different PEO molecular weights for Fe3O4@SiO2-PEO-mal nanoparticles were used for characterizing of MBP-Hep I immobilization. The results showed that all four kinds of Fe3O4@SiO2-PEO-mal magnetic nanoparticles (6k, 20k, 35k and 100k for PEO) exhibited excellent adsorption capacities and the adsorption ratio increased as the PEO molecular weight increased from 6k to 100k. All four kinds of immobilized MBP-Hep I exhibited significantly improved stability at 30 °C compared with free MBP-Hep I and their half-lives were 20–50 times that of the free MBP-Hep I. Fe3O4@SiO2-PEO-mal nanoparticles with a PEO molecular weight of 100k were best able to immobilize MBP-Hep I (Fe3O4@SiO2-PEO100k-mal-MBP-Hep I). The molecular weight distribution profiles and anticoagulant activities, obtained from heparin depolymerization by free Hep I, free MBP-Hep I and Fe3O4@SiO2-PEO100k-mal-MBP-Hep I were the same. Furthermore, Fe3O4@SiO2-PEO100k-mal-MBP-Hep I exhibited reasonable reusability during enzymatic production of low molecular weight heparins (LMWHs). 相似文献
7.
A RNase has been partially purified from rat reticulocytes induced by phenylhydrazine. This enzyme has an optimum pH of 7.5 and degrades RNA endonucleolytically as evidenced by the analysis of degradation products. The activity is destroyed by heat treatment (pH 6.5, 80 °, 5 min). Many metal ions are inhibitory for the activity. The enzyme was inactivated almost completely by 0.5 mm HgCl2. Monovalent ions, including Nad, KCl, NH4Cl, and (NH4)2SO4, inhibit the enzyme by about 90% at concentrations of 0.1–0.2 m. The molecular weight of this enzyme is about 16,000 as determined by gel filtration. A latent RNase with higher molecular weight is present in the crude extract of the cells. 相似文献
8.
Proteins synthesized by soma located in L4 dorsal root ganglia and supplied to the axonal branches extending centrally in the dorsal root and peripherally towards the sciatic nerve were analyzed for radioactivity following injections of [3H] leucine into the L4 dorsal root ganglia. All proteins located in the dorsal root and sciatic nerve were analyzed by SDS acrylamide gel electrophoresis at various times post injection. The differences in radioactivity between the dorsal root and sciatic nerve proteins were mainly quantitative and not qualitative, with many proteins of various molecular weight ranges being transported into both segments. Generally, it appears that in both axonal branches the high molecular weight proteins are transported at the highest rate, medium weights slower and low molecular weight proteins slowest. More proteins of high and low molecular weights are transported into the dorsal root whereas more of those of medium molecular weight are transported towards the sciatic nerve. 相似文献
9.
10.
Ivana Vančurová Aleš Vančura Jindřich Volc Jiří Neužzil Miroslav Flieger Gabriela Basařová Vladislav Běhal 《Archives of microbiology》1988,150(5):438-440
Alanine dehydrogenase was purified to near homogeneity from cell-free extract of Streptomyces aureofaciens, which produces tetracycline. The molecular weight of the enzyme determined by size-exclusion high-performance liquid chromatography was 395 000. The molecular weight determined by sodium dodecyl sulfate gel electrophoresis was 48 000, indicating that the enzyme consists of eight subunits with similar molecular weight. The isoelectric point of alanine dehydrogenase is 6.7. The pH optimum is 10.0 for oxidative deamination of L-alanine and 8.5 for reductive amination of pyruvate. K
M values were 5.0 mM for L-alanine and 0.11 mM for NAD+. K
M values for reductive amination were 0.56 mM for pyruvate, 0.029 mM for NADH and 6.67 mM for NH4Cl.Abbreviation AlaDH
alanine dehydrogenase 相似文献
11.
J.F. Evans P. Dupuis A.W. Ford-Hutchinson 《Biochimica et Biophysica Acta (BBA)/General Subjects》1985,840(1):43-50
Leukotriene A4 hydrolase was rapidly and extensively purified from rat neutrophils using anion exchange and gel filtration high-pressure liquid chromatography. The enzyme which converts the allylic epoxide leukotriene A4 to the 5,12-dihydroxyeicosatetraenoic acid leukotriene B4 was localized in the cytosolic fraction and exhibited an optimum activity at pH 7.8 and apparent Km for leukotriene A4 between 2 · 10?5 and 3 · 10?5 M. The purified leukotriene A4 hydrolase was shown to have a molecular weight of 68 000 on sodium dodecylsulfate polyacrylamide gel electrophoresis and of 50 000 by gel filtration. The molecular weight and monomeric native form of this enzyme are unique characteristics which distinguish leukotriene A4 hydrolase from previously purified epoxide hydrolases. 相似文献
12.
Aggarwal S Tandon CD Forouzandeh M Singla SK Kiran R Jethi RK 《Molecular and cellular biochemistry》2000,210(1-2):109-119
Human renal calculi surgically removed from kidney stone patients were obtained and chemically analysed. Stones with CaOx (calcium oxalate) as the major component were washed in 0.15 M NaCl with gentle stirring for 48 h and then pulverised to a fine powder. The powder was extracted with 0.05 M EGTA, 1 mM PMSF and 1% - mercaptoethanol for 4 days at 4°C, the suspensions and the supernatants obtained were filtered through an Amicon Model 200 apparatus (mol. wt. cut off of 10,000 daltons) under nitrogen at 40 p.s.i. and concentrated to a known volume. The method of Nakagawa et al. [7] was employed to study the ability of > 10 kDa fractions to influence COM growth using metastable solution of CaCl2 and Na2C2O2 containing traces of 14C-oxalic acid. Potent biomolecules having the ability to influence CaOx precipitation were subjected to isolation, purification and characterization. Standard biochemical procedures, e.g. ultracentrifugation, ion-exchange chromatography, molecular sieve chromatography and SDS-PAGE, etc., were employed. Results revealed that human renal calculi extract contains biomolecules that can inhibit as well as stimulate the growth of preformed COM (calcium oxalate monohydrate) crystals. Most potent stimulator of CaOx growth was found to have a molecular weight of 66 kDa. 相似文献
13.
Purification and Partial Characterization of a Genetically-Defined Superoxide Dismutase (SOD-1) Associated with Maize Chloroplasts 总被引:7,自引:1,他引:6
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The chloroplast-associated form of superoxide dismutase from maize (Zea mays L.) (SOD-1) has been purified by a stepwise procedure consisting of (NH4)2SO4 fractionation, G-100 Sephadex gel filtration, DEAE-Sephacel chromatography, and hydroxylapatite chromatography. This procedure resulted in a single band on sodium dodecyl sulfate-polyacrylamide gels indicating that the preparation is homogeneous. The holoenzyme molecular weight was estimated at 31,000 to 33,000 by gel filtration. The subunit molecular weight of this dimeric protein was estimated at 14,500 on sodium dodecyl sulfate-polyacrylamide gels. Studies involving amino acid composition analysis, immunological cross-reactivity, in vitro subunit hybridizations, and H2O2 sensitivity indicate that SOD-1 differs significantly from SOD-2 and SOD-4, the other cupro-zinc forms of SOD from maize. The possible physiological role of SOD-1 within the chloroplast is discussed. 相似文献
14.
A series of pullulan fractions with molecular weights in the range 5 × 103 to 8 × 105 were prepared. The weight-average molecular weight (Mw) of all the samples was determined by sedimentation equilibrium. The hydrodynamic properties of pullulan in aqueous solution were investigated by viscometry and ultracentrifugation. The experimental results indicate that pullulan molecules in water are fairly stable and behave as expanded random coils when Mw is above 2 × 104. The molecular weight distributions of the fractions were measured by gel filtration. The ratio Mw/Mn was close to 1·1, except for a sample with the highest Mw.It is concluded that the pullulan fractions prepared by the present work are well characterized and have a narrow molecular weight distribution. They may be useful as standard samples for studies of water-soluble polymers. 相似文献
15.
Calf thymus and salmon sperm deoxyribouncleie acid were degraded by high-shear stirring to molecular weights M in the range of 1.3–3.2 × 106 and purified by chromatography on methylated bovine serum albumin. Dynamic viscoelastic properties of the fragmented products, in aqueous glycerol solutions in the concentration range of c = 0.003–0.01 g./ml., were investigated with the apparatus of Birnboim and Ferry. At values of the product cM higher than 4 × 103, the frequency dependence of the components of the complex shear modulus, G′ and G″, displayed a plateau region in which G′ > G″ – ων1ηS, similar to that observed in concentrated solutions of coiling polymers where it is attributed to an entanglement network (ω is radian frequency, ν1 volume fraction of solvent, and η8, solvent viscosity). The width of this plateau region on the logarithmic frequency scale is given by Δ = 3.8 (log cM – 3.56). At lower values of cM, the frequency dependence is intermediate between those predicted by the theory of Zimm for flexible coiled macromolecules and by the theory of Kirkwood and Auer for rods. Fitting to the Zimm theory gives highly discrepant values for molecular weights, while fitting the low-frequency end of the dispersion to the Kirkwood-Auer theory gives reasonable agreement for both molecular weight and rotary diffusion coefficient. It is concluded that the helical fragments appear as nearly rigid rods in their behavior at very low frequencies, but at higher frequencies reveal substantial bending flexibility. 相似文献
16.
Mareková M Lauková A Skaugen M Nes I 《Journal of industrial microbiology & biotechnology》2007,34(8):533-537
The new bacteriocin, termed enterocin M, produced by Enterococcus faecium AL 41 showed a wide spectrum of inhibitory activity against the indicator organisms from different sources. It was purified
by (NH4)2SO4 precipitation, cation-exchange chromatography and reverse phase chromatography (FPLC). The purified peptide was sequenced
by N-terminal amino acid Edman degradation and a mass spectrometry analysis was performed. By combining the data obtained
from amino acid sequence (39 N-terminal amino acid residues was determined) and the molecular weight (determined to be 4 628 Da)
it was concluded that the purified enterocin M is a new bacteriocin, which is very similar to enterocin P. However, its molecular
weight is different from enterocin P (4 701.25). Of the first 39 N-terminal residues of enterocin M, valine was found in position
20 and a lysine in position 35, while enterocin P has tryptophane residues in these positions. 相似文献
17.
Characterization of the octamer of histones free in solution 总被引:15,自引:0,他引:15
The nucleosome “core protein” isolated from chromatin in high-salt solutions (2 m-NaCl) has been characterized in detail. The preparation described yields material which is stable for prolonged periods at either 4 °C or 37 °C. It has an apparent partial specific volume of 0.767 ml/g and a sedimentation coefficient (S20,w0) of 4.77 (±0.04) S. The molecular weight, determined by sedimentation equilibrium, is 107,500 (±7700), which is compatible with an octameric structure of the form (H2A)2(H2B)2(H3)2(H4)2, as previously proposed.The sedimentation coefficient and molecular weight are very similar to those of cross-linked core protein, which is known to be octameric. 相似文献
18.
Alexandre Rothen 《The Journal of general physiology》1940,24(2):203-211
Electrophoretic studies on purified crystalline ribonuclease showed the absence of any impurities differing in mobility from the bulk of material. The isoelectric point of ribonuclease was found by electrophoresis to be at about pH 7.8. Ultracentrifuge studies indicated fair homogeneity of ribonuclease in solution. Only one moving component has been observed. The molecular weight of ribonuclease was found to be 12,700 from rate of sedimentation (S
25 = 1.85 x 10–13 in 0.5 M (NH4)2SO4) and diffusion measurement (D = 1.36 x 10–6 in 0.5 M (NH4)2SO4), in good agreement with the average value of 13,000 found from equilibrium measurements. This low value for the molecular weight of a protein would seem to discredit the value 17,600 as representing a universal unit weight for proteins in general. 相似文献
19.
Human Serum Cholinesterase,a Tetramer 总被引:3,自引:0,他引:3
AFTER electrophoresis of human serum in starch gel1, there are at least four bands of Cholinesterase activity. Most of the enzyme is concentrated in the band with least mobility (C4) while three faster moving bands (C1, C2, C3) are of lower molecular weight as shown by gel filtration2. Although electrophoresis in Polyacrylamide reveals additional bands3, all seem to be a single genetic entity as shown by experiments on persons homozygous for the silent gene4,5. The C4 enzyme has a molecular weight of at least 250,000 (ref. 6) and can be assumed to be composed of more than one polypeptide chain. By preparing an enzyme of altered mobility and forming hybrids of it with the usual enzyme, we have obtained results that suggest that Cholinesterase is a tetramer. 相似文献
20.
The molecular weight of rhodopsin and the nature of the rhodopsin-digitonin complex 总被引:5,自引:0,他引:5
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HUBBARD R 《The Journal of general physiology》1954,37(3):381-399
The sedimentation behavior of aqueous solutions of digitonin and of cattle rhodopsin in digitonin has been examined in the ultracentrifuge. In confirmation of earlier work, digitonin was found to sediment as a micelle (D-1) with an s20 of about 6.35 Svedberg units, and containing at least 60 molecules. The rhodopsin solutions sediment as a stoichiometric complex of rhodopsin with digitonin (RD-1) with an s20 of about 9.77 Svedberg units. The s20 of the RD-1 micelle is constant between pH 6.3 and 9.6, and in the presence of excess digitonin. RD-1 travels as a single boundary also in the electrophoresis apparatus at pH 8.5, and on filter paper at pH 8.0. The molecular weight of the RD-1 micelle lies between 260,000 and 290,000. Of this, only about 40,000 gm. are due to rhodopsin; the rest is digitonin (180 to 200 moles). Comparison of the relative concentrations of RD-1 and retinene in solutions of rhodopsin-digitonin shows that RD-1 contains only one retinene equivalent. It can therefore contain only one molecule of rhodopsin with a molecular weight of about 40,000. Cattle rhodopsin therefore contains only one chromophore consisting of a single molecule of retinene. It is likely that frog rhodopsin has a similar molecular weight and also contains only one chromophore per molecule. The molar extinction coefficient of rhodopsin is therefore identical with the extinction coefficient per mole of retinene (40,600 cm.2 per mole) and the E(1 per cent, 1 cm., 500 mµ) has a value of about 10. Rhodopsin constitutes about 14 per cent of the dry weight, and 3.7 per cent of the wet weight of cattle outer limbs. This corresponds to about 4.2 x 106 molecules of rhodopsin per outer limb. The rhodopsin content of frog outer limbs is considerably higher: about 35 per cent of the dry weight, and 10 per cent of the wet weight, corresponding to about 2.1 x 109 molecules per outer limb. Thus the frog outer limb contains about five hundred times as much rhodopsin as the cattle outer limb. But the relative volumes of these structures are such that the ratio of concentrations is only about 2.5 to 1 on a weight basis. Rhodopsin accounts for at least one-fifth of the total protein of the cattle outer limb; for the frog, this value must be higher. The extinction (K500) along its axis is about 0.037 cm.2 for the cattle outer limb, and about 0.50 cm.2 for the frog outer limb. 相似文献