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1.
W J Strittmatter F Hirata J Axelrod 《Biochemical and biophysical research communications》1979,88(1):147-153
Ca2+-ATPase activity in human erythrocytes is increased by the enzymatic methylation of membrane phospholipids. Erythrocyte membranes incubated in the presence of the methyl donor, S-adenosyl-L-methionine, demonstrate increased Ca2+ stimulated ATP hydrolysis, increased [45Ca2+] efflux from erythrocyte ghosts and synthesis of phosphatidyl-N-monomethylethanolamine. The increase in Ca2+-ATPase activity is due to an increase in Vmax, and not due to changes in affinity for ATP or Ca2+. The concentration of S-adenosyl-L-methionine needed to stimulate Ca2+-ATPase closely matches that needed for the methylation of phosphatidylethanolamine. Both the stimulation of Ca2+-ATPase and the methylation of phospholipids are inhibited by the methyltransferase inhibitor, S-adenosyl-L-homocysteine. Membrane fluidity is increased by phospholipid methylation, which may be the mechanism for Ca2+-ATPase stimulation. 相似文献
2.
V Niggli P Ronner E Carafoli J T Penniston 《Archives of biochemistry and biophysics》1979,198(1):124-130
The stimulation of the (Ca2+ + Mg2+)ATPase of erythrocyte ghosts by calmodulin was observed not only in intact ghosts, but also in the solubilized (Triton X-100) and partially purified, reconstituted (phosphatidylserine liposomes) forms. Since the solubilized form of the enzyme migrated on Sepharose 6B at a position corresponding to a molecular weight of about 150,000, these results show that calmodulin stimulates by direct interaction with the ATPase complex. Additionally, the effects of calmodulin on erythrocyte ghosts prepared by the Dodge-EDTA method (hypotonic ghosts) and by the method of Ronner et al. (involving lysis followed by an isotonic wash repeated several times) were compared (P. Ronner, P. Gazzotti, and E. Carafoli, 1977, Arch. Biochem. Biophys. 179, 578–583). The (Ca2+ + Mg2+)ATPase of the hypotonic ghosts was low and was stimulated by added calmodulin while that of the isotonic ghosts was high and changed only slightly upon calmodulin addition; this difference in response to calmodulin persisted in the solubilized and reconstituted forms. Hypotonic ghosts bound 125I-labeled calmodulin, while isotonic ghosts did not. This comparison of two types of ghosts showed that isotonic ghosts possess an intact calmodulin-(Ca2+ + Mg2+)ATPase complex, and that the calmodulin remained with the ATPase during solubilization and reconstitution. The isotonic preparation is a particularly useful method of preparing ghosts with an intact calmodulin-ATPase complex, since it requires no special equipment and produces an enzyme activity which is stable to freezing. 相似文献
3.
Eugene E. Quist 《Biochemical and biophysical research communications》1980,92(2):631-637
In the presence of 1.0 mM ATP and MgCl2, the specific viscosity of suspensions of human erythrocyte ghosts decreases 35% in 20 minutes at 22°C. The changes in viscosity are a sensitive index of Mg-ATP dependent shape changes in these membranes. Low concentrations of Ca2+ (1 to 5 μM) inhibit Mg-ATP dependent viscosity changes. If ghosts were preincubated with 1 mM Mg-ATP and 20 μM A23187 to produce a maximal decrease in viscosity, addition of 10 μM Ca2+ to the preincubated ghosts increased the viscosity to levels observed in ghosts preincubated without ATP. Ca2+ (1 to 5 μM) also inhibited Mg2+ dependent phosphorylation 30% and stimulated dephosphorylation 25% in ghost membranes. These effects of Ca2+ on viscosity and phosphorylation may be due to a membrane bound Ca2+ phosphatase activity which dephosphorylates membranes phosphorylated by a Mg2+ dependent kinase activity. 相似文献
4.
In Dictyostelium discoideum, the formation of multicellular aggregates represents the first morphogenetic event that leads ultimately to the construction of fruiting bodies. The altered adhesive properties of the cells can be demonstrated in ghosts derived from them which consist of largely intact membranes containing a few empty vesicles and exploded mitochondria but lacking nuclei, RNA, soluble cytoplasm and ATP [4]. A cofactor requirement for the aggregation of the ghosts can be satisfied by the following divalent cations: Ca2+, Mn2+, Zn2+ and Cu2+. In this paper it is shown that associated with the acquisition of aggregative competence is a 15–20-fold increase in the ghosts of sites capable of binding either Ca2+ or Mn2+ with relatively high affinity. 相似文献
5.
Manganous Ion as a Spin Label in Studies of Mitochondrial Uptake of Manganese 总被引:4,自引:0,他引:4
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Manganous ion (Mn2+) has been used as a spin label for studies of divalent cation uptake by rat liver mitochondria. Spin exchange, observed in the electron paramagnetic resonance (EPR) spectrum of a fraction of the transported Mn2+, shows that this fraction is bound in regions of high local concentration within the mitochondria. The average separation of manganese ions in that fraction is estimated to be 4.0 ±0.6 A at the time of greatest concentration. 相似文献
6.
Proton and phosphorus-31 nuclear spin–lattice relaxation times T1 and spin–spin relaxation times T2 have been measured on the single-stranded polyriboadenylic acid [poly(A)]–Mn2+ system in a neutral D2O solution in the temperature range 10°–90°C at 100 and 40.5 MHz, respectively, with the Fourier transform nmr method. Minimum values of T1 have been found for all these nuclei, which have enabled the exact estimation of apparent distances from Mn2+ to H2, H8, H1′, and the phosphorus nucleus to be 4.7, 4.1, 5.2, and 3.0 Å, respectively. The electron spin of Mn2+ penetrates into the phosphorus nucleus, giving 31P hyperfine coupling of more than 106 Hz. Evidence of penetration of the electron spin into H8 and H2 is also obtained, suggesting direct coordination of nitrogen atoms of the adenine ring to the Mn2+ Ion. Combined with the result from proton relaxation enhancement of water, it is concluded that every Mn2+ ion added is bound directly to two phosphate groups with a Mn2+–phosphorus distance of 3.3 Å, while a part of the Mn2+ ions are simultaneouly bound to the adenine ring. It is estimated that 39 ± 13% and 13 ± 5% of Mn2+ are coordinated by N7 and N3 (or N1), respectively. The motional freedom of poly(A) in the environment of the Mn2+ binding site has been found to be quenched to the extent that the rotational motion becomes several times slower than that of the corresponding Mn2+–free poly(A). The activation energies for the molecular motion are, however, practically unchanged from those for Mn2+–free poly(A), and are found to be 8.3, 8.5, 6.1, and 8.7 kcal/mol for H8, H2, H1′, and phosphorus, respectively. T2 of phosphorus is determined by the dissociation rate (k?1) of Mn2+ from the phosphate group for the whole temperature range studied with activation enthalpy of 6.5 kcal/mol. The dissociation rates of Mn2+ from the adenine ring are also estimated from proton T2 values below 50°C. 相似文献
7.
When human red cells are incubated at 37°C with the artificial electron donor system ascorbate + phenazine methosulphate the fluxes of Rb+ (K+) through the cell membrane are increased. The effect of this donor system is much stronger in energy-depleted than in normal cells. The same effects are produced by HS-glutathione, NADH or NADPH loaded into resealed ghosts, but these electron donors were ineffective when added to the incubation medium. The Rb+ (K+) fluxes induced by electron donors resemble closely those induced by an increase of intracellular Ca2+ (Gardos effect). The electron donors require the presence of intracellular Ca2+ to be effective, but at levels that do not stimulate by themselves the fluxes of K+. Flavoenzyme inhibitors (atebrin and chlorpromazine), oligomycin and quinine prevented the effects of both electron donors and Ca2+ alone; antimycin, uncouplers and ethacrynic acid inhibited them partially; ouabain, furosemide, and rotenone had no effect.The results could be explained if the effect of electron donors is to bring about a change in the redox state of some membrane component(s) that makes intracellular Ca2+ more effective to elicit rapid K+ movements. Plasma membrane oxidoreductase activities could be engaged in this change. 相似文献
8.
The electron spin resonance (ESR) lines of nitroxide spin labels are broadened by electron spin exchange reactions that take place during collisions with paramagnetic ions. The degree of line broadening is greatly reduced when the paramagnetic ion forms a coordination bond with certain functional groups on organic molecules. These observations form the basis for a spin-label assay for metal ion chelation and complex formation. This paper describes the characteristics of such an assay for divalent nickel ions and the spin label TEMPONE (2,2,6,6-tetramethylpiperidone-N-oxyl). The chelation of Ni2+ by cysteine and the interaction of Ni2+ with sodium dodecyl sulfate micelles and phospholipid vesicles are demonstrated. In addition to monitoring interactions of paramagnetic ions, the assay also allows the detection of interactions of nonparamagnetic ions that compete with the paramagnetic ions for binding sites. A kinetic analysis of competition between Ni2+ and Zn2+ ions for binding sites on phospholipid vesicles is presented. There are several advantages of the spin-label line-broadening assay compared to other conventional assays for metal chelation and complex formation. The line-broadening assay does not require that the sample be optically clear or chemically defined, it requires only very small quantities of material, it can detect as little as 0.4 to 1 μmol of complexing agent, and it may be utilized in complex biological systems including subcellular organelles and macromolecules. 相似文献
9.
Donald E. Riley John M. Keller 《Biochimica et Biophysica Acta (BBA)/General Subjects》1976,444(3):899-911
The polypeptide species of non-membranous nuclear ghosts from purified cell nuclei are conserved among a variety of human, hamster and mouse cell types studied, including HeLa, BHK, 3T6 and Hep-2 cell lines. The polypeptide species present in nuclear ghosts from HeLa cells synchronized in various stages of the cell cycle are largely the same with minor variations. The isolated nuclear ghosts are similar, in terms of polypeptide composition, to other residual nuclear structures isolated by independent techniques. The nuclear ghosts appear as flattened sac-like structures when viewed by scanning electron microscopy. Transmission electron microscopy of the nuclear ghosts reveals ring-like structures which may represent the nuclear pores. Also observed are novel rod-shaped structures approximately 260 nm in length and 50 nm in diameter. The latter images either arise by a rearrangement during isolation of the nuclear ghost macromolecules or are a heretofore undescribed structure of intact nuclei. 相似文献
10.
JoséD. Cavieres 《生物化学与生物物理学报:生物膜》1984,771(2):241-244
An average target size of 251 kDa has been obtained for the (Ca2+ + Mg2+)-ATPase of calmodulin-depleted erythrocyte ghosts by radiation inactivation with 16 MeV electrons. This is close to twice the size of the purified calcium-pump polypeptide. When calmodulin was included during the ATPase assay, a component of about 1 MDa appeared in addition to the activated dimer. 相似文献
11.
A detailed study has been made of the permeability characteristics of human erythrocyte ghosts prepared under isoionic conditions by a glycol-induced lysis (Billah, M.M., Finean, J.B., Coleman, R. and Michell, R.H. (1976) Biochim. Biophys. Acta 433, 45–54). Impermeability to large molecules such as dextran (average molecular weight 70 000) was restored immediately and spontaneously after each of the 5–7 lyses that were required to remove all of the haemoglobin. Permeabilities to smaller molecules such as MgATP2?, [3H]inositol and [14C]choline were initially high but could be greatly reduced by incubation at 37°C for an hour. The extent of such resealing decreased as the number of lyses to which the ghosts had been subjected increased. Both removal of haemoglobin and permeabilities to small molecules were affected significantly by pH, Ca2+ concentrations and divalent cation chelators. Maximum resealing was achieved in ghosts prepared in the basic ionic medium (130 mM KCl, 10 mM NaCl, 2 mM MgCl2, 10 mM acid (HEPES)) at pH 7.0 (0°C) and with a calcium level around 10?5 M. Acidic pH facilitated the removal of haemoglobin whilst the presence of divalent cation chelators slowed down its release. Retention of K+ by ghosts loaded with K+ during the first lysis and subsequently incubated at 37° C was substantial but little K+ could be retained within the haemoglobin-free ghosts. Permeability of the ghosts to K+ after one lysis was affected by temperature, pH, Ca2+ concentrations and by the presence of divalent cation chelators. 相似文献
12.
Bulk water transport in reconstituted ghosts is statistically comparable to that in the parent red cells, and is unaffected by incorporation of Ca2+ over the range of 0.01 to 1 mM. Brief exposure of ghosts to p-chloromercuribenzene sulfonate results in a supression of osmotic water flow but leaves K+ permeability unchanged. Incorporation of p-chloromercuribenzene sulfonate provokes extremely rapid K+ loss which can be counteracted by simultaneous inclusion of Ca2+.Erythrocyte ghosts, when prepared with a small amount of Ca2+, demonstrate recovery of normal impermeability to choline, sucrose, Na+ and inulin and have an improved K+ retention over Ca2+-free preparations.The rate of passive transport of K+ from unwashed erythrocyte ghosts was measured during the initial few minutes of efflux. The initial rates vary in a bimodal fashion with the concentration of Ca2+ incorporated at the time of hemolysis. In low concentrations (0.01–0.1 mM), Ca2+ protects the K+ barrier while at higher concentrations (0.1–1.0 mM) it provokes a K+ leakage ranging from 7 to 50 times the normal rate of passive K+ loss. The Ca2+-induced K+ leak is thus a graded response rather than a discrete membrane transport state. The transition from a Ca2+-protected to a Ca2+-damaged membrane occurs upon an increase in Ca2+ concentration of less than 50 μmoles/l. 相似文献
13.
Buchachenko AL Kouznetsov DA Arkhangelsky SE Orlova MA Markarian AA 《Cell biochemistry and biophysics》2005,43(2):243-251
The rates of adenosine triphosphate (ATP) production by isolated mitochondria and mitochondrial creatime kinase incubated
in isotopically pure media containing, separately, 24Mg2+, 25Mg2+, and 26Mg2+ ions were shown to be strongly dependent on the magnesium nuclear spin and magnetic moment. The rate of adenosine 5′-diphosphate
phosphorylation in mitochondria with magnetic nuclei25Mg is about twice higher than that with the spinless, nonmagnetic nuclei24.26Mg. When mitochondrial oxidative phosphorylation was selectively blocked by treatment with 1-methylnicotine amide, 25Mg2+ ions were shown to be nearly four times more active in mitochondrial ATP synthesis than 24,26Mg2+ ions. The rate of ATP production associated with creatine kinase is twice higher for 25Mg2+ than for 24.26Mg and does not depend on the blockade of oxidative phosphorylation. There is no difference between 24Mg2+ and 26Mg2+ effects in both oxidative and substrate phophorylation. These observations demonstrate that the enzymatic phosphorylation
is a nuclear spin selective process controlled by magnetic isotope effect. The reaction mechanism proposed includes a participation
of intermediate ion-radical pairs with Mg+ cation as a radical partner. Therefore, the key mitochondrial phosphotransferases work as a magnesium nuclear spin mediated
molecular machines. 相似文献
14.
Toshio Iwasaki Reinhard Kappl Gerhard Bracic Nobutaka Shimizu Daijiro Ohmori Takashi Kumasaka 《Journal of biological inorganic chemistry》2011,16(6):923-935
The crystal structure of the ISC-like [2Fe–2S] ferredoxin (FdxB), probably involved in the de novo iron-sulfur cluster biosynthesis (ISC) system of Pseudomonas putida JCM 20004, was determined at 1.90-Å resolution and displayed a novel tail-to-tail dimeric form. P. putida FdxB lacks the consensus free cysteine usually present near the cluster of ISC-like ferredoxins, indicating its primarily electron transfer role in the iron-sulfur cluster. Orientation-selective electron–nuclear double resonance spectroscopic analysis of reduced FdxB in conjunction with the crystal structure has identified the innermost Fe2 site with a high positive spin population as the nonreducible iron retaining the Fe3+ valence and the outermost Fe1 site as the reduced iron with a low negative spin density. The average g max direction is skewed, forming an angle of about 27.3° (±4°) with the normal of the [2Fe–2S] plane, whereas the g int and g min directions are distributed in the cluster plane, presumably tilted by the same angle with respect to this plane. These results are related to those for other [2Fe–2S] proteins in different electron transport chains (e.g. adrenodoxin) and suggest a significant distortion of the electronic structure of the reduced [2Fe–2S] cluster under the influence of the protein environment around each iron site in general. 相似文献
15.
The apo protein of imidazole glycerol phosphate dehydratase (IGPD) from Saccharomyces cerevisiae combines stoichiometrically with certain specific divalent metal cations to assemble the catalytically active form comprising
24 protein subunits and tightly bound metal. VO2+ ions react similarly but, uniquely, result in a metallo-protein (VO-IGPD) with neither catalytic activity nor the ability
to bind to the reaction intermediate analogue, 2-hydroxy-3-(1,2,4-triazol-1-yl) propylphosphonate. Since VO2+ apparently assembles the quaternary structure correctly, it is used in the present study as a spin probe to investigate the
metal centre coordination environment by electron paramagnetic resonance (EPR) and electron nuclear double resonance (ENDOR)
spectroscopy. At neutral pH, the EPR spectrum of VO-IGPD reveals at least three distinct VO2+ sub-spectra with one predominant at low pH. The spin Hamiltonian parameters for some of the sub-spectra are consistent with
51V having nitrogen in the inner-sphere equatorial coordination environment from, most probably, multiple coordinating histidines.
Further evidence for inner-sphere nitrogen ligands is obtained from ENDOR spectroscopy. The spectra of the low rf region show
signals from interactions with 14N which are consistent with couplings to the imino nitrogen of coordinated histidine residues. In addition a number of proton
ENDOR line pairs are resolved. Of the few that disappear upon exchange of the protein into D2O, one most likely originates from the exchangeable proton of the N-H group of a coordinated histidine imidazole. 1H-ENDOR line pairs from non-exchangeable protons with splittings of approximately 3 MHz can be attributed to imidazole carbon
protons. Thus, most of the couplings observed by ENDOR are consistent with being from the imidazole heterocycle of one or
more histidine ligands.
Received: 27 June 1996 / Accepted: 14 March 1997 相似文献
16.
To help interpret the electron spin resonance (esr) spectra of spin-labeled actin, the positions of attachment of the spin labels, N-(1-oxyl-2,2,6,6-tetramethyl-4-piperidinyl) maleimide and N-(1-oxyl-2,2,6,6-tetramethyl-4-piperidinyl) iodoacetamide to rabbit skeletal muscle actin have been determined. For this purpose spin-labeled peptides released by tryptic digestion of the spin-labeled actin were isolated by chromatography and identified from their positions of elution and amino acid composition.With purified F-actin that had not undergone structural changes both labels reacted exclusively with the sulfhydryl group of the C-terminal sequence. But if the actin was stored in the F-form in the absence of ATP it evidently underwent a structural alteration because reaction was then at another sulfhydryl group, in the N-terminal sequence, and the actin had an irregular appearance in the electron microscope. ADP and tripolyphosphate were as effective as ATP in preventing this alteration. A maximum of 1 equiv of spin label was bound, irrespective of the site of labeling, and the two sites appeared to be mutually exclusive, possibly because they are adjacent. With G-actin, and with actin denatured by guanidine hydrochloride, there was also reaction at other sites. The shapes of the esr spectra of F-actin that contained Mg2+, Ca2+, or Mn2+ did not depend on whether labeling was at the C- or N-terminal positions, although F-actin labeled in the latter position contained a small proportion of highly mobile label, possibly a result of denaturation. The reduction in the size of the esr signal of labeled G-actin on replacing Mg2+ with Mn2+ did not appear to be dependent on the position of labeling. 相似文献
17.
Lipid composition of envelopes,prolamellar bodies and other plastid membranes in etiolated,green and greening wheat leaves 总被引:1,自引:1,他引:0
Summary Comparative studies of lipid composition were made on prolamellar bodies, envelopes and other plastid membranes separately extracted from etiolated, green or greening (intermittent or continuous light) wheat (Triticum sativum L.) leaves. The different membrane fractions were examined by electron microscopy.The major lipid was digalactosyldiglyceride in the envelopes and prolamellar bodies and monogalactosyldiglyceride in stroma lamellae and grana. Phosphatidylcholine represented 60% of total phospholipids in the envelopes, 30% in prolamellar bodies and 14% in grana. All types of envelopes had the same lipid proportions.For all lipids the lowest fatty acid unsaturation was always found in the envelope membranes. The relative amount of {ie193-1} acid in the phosphatidylglycerol of envelopes increased from 4% (etioplasts) to an average of 15% (etiochloroplasts and chloroplasts).Abbreviations DGDG
digalactosyldiglyceride
- MGDG
monogalactosyldiglyceride
- PC
phosphatidylcholine
- PE
phosphatidylethanolamine
- PG
phosphatidylglycerol
- PI
phosphatidylinositol
- PS
phosphatidylserine
- SL
sulfolipid 相似文献
18.
Summary Different cytochemical methods were employed to demonstrate the existence of Ca2+-binding sites (Ca2+-bs) at the membranes of barley root tip cells, involving addition of CaCl2 (10 mM or 1 mM) to all aqueous solutions used for tissue processing for electron microscopy, treatment of ultrathin sections by Ca-chelating agents, enzymic digestion of ultrathin sections and modification of Wachstein-Meisel procedure for localization of Ca2+-dependent ATPase activity. Addition of 10 mM CaCl2 to the fixatives and rinsing solutions causes electron-dense globules (EDG) to be formed in a variety of cells, those in cortical cells being associated mainly with the plasma membranes, in root cap cells with the plasmalemma as well as with majority of intracellular membranes. The obligatory presence of EDG at the membranes of Golgi vesicles and secretory vesicles approaching plasmalemma was revealed in the secreting root cap cells. Besides, electron opaque connecting material was found between the plasmalemma and adjacent secretory vesicle membranes. In true meristematic cells Ca-supplemented solutions induce formation of EDG localized at the ER membranes, and nuclear and plastid envelopes. In root cells of seeds germinated in the presence of 1 mM CaCl2 electron opaque deposits were found only in local areas of plasmalemma collars around plasmodesmata neck regions, contacting the terminals of subsurface ER channels. In control speciemens (germination, fixation and washing without added CaCl2) EDG were absent in cortical and ground meristem cells, but present in root cap cells, although their number and average size were greatly reduced.Treatment of thin sections by 10mM EGTA or EDTA led to complete removing of EDGs, electron-transparent holes replacing them. Digestion by a variety of proteolytic enzymes and by phospholipase A induced partial destruction of EDG matrices, confirming the presence of protein as well as of phospholipid membrane components. Visualization of electron-dense granular product of cytochemical Ca-ATPase reaction at the same membrane areas where EDG were located suggests that one of the Ca-binding proteins in EDG may represent Ca-ATPase.It is proposed that EDG at plant cell membranes have a certain resemblance to the Ca2+-bs revealed by the same method on plasma membrane of a variety of animal cells. The data obtained are discussed regarding possible regulatory roles of calcium ions in plant cells, especially in exocytotic secretion. 相似文献
19.
The role of a transmembrane Ca2+ gradient in anion transport by Band 3 of human resealed erythrocyte ghosts has been studied. The results show that a transmembrane Ca2+ gradient is essential for the conformation of erythrocyte Band 3 with higher anion transport activity. The dissipation of the transmembrane Ca2+ gradient by the ionophore A23187 inhibits the anion transport activity. The extent of this inhibition approaches 90% as the Ca2+ concentration on both sides of the ghost membrane is increased to 1.0 mM and half-maximum inhibition is observed at 0.25 mM Ca2+. Addition of ATP (0.4 mM) to the resealing medium can partly reestablish the transmembrane Ca2+ gradient by activation of Ca2+-ATPase and alleviate the inhibition to some extent. N-ethylmaleimide, an inhibitor of erythrocyte Ca2+-ATPase, prevents such restoration. Electron micrographs reveal that numerous larger intramembranous particles can be observed on the P-faces of freeze-fractured resealed ghosts in the absence of a transmembrane Ca2+ gradient.Abbreviations DPA
dipicolinic acid
- EITC
eosin 5-isothiocyanate
- DIDS
4,4-diisothiocyanostilbene-2,2-disulfonate
- TES
N-Tris-(hydroxymethyl)methyl-2-aminoethane sulfonic acid
- PMSF
phenylmethyl-sulfonylfluoride
- NEM
N-ethylamaleimide
- BSA
bovine serum albumin
- EGTA
ethyleneglycol-bis (aminoethylether)-tetra-acetic acid
- EITC-Band 3
Band 3 labeled with EITC
- Cai
Ca2+ inside resealed ghosts
- Cao
Ca2+ outside resealed ghosts 相似文献
20.
Expansion and apparent fluidity decrease of nuclear membranes induced by low Ca/Mg. Modulation of nuclear membrane lipid fluidity by the membrane-associated nuclear matrix proteins? 总被引:1,自引:0,他引:1
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《The Journal of cell biology》1978,79(2):479-490
Macronuclei isolated from Tetrahymena are contracted in form (average diameter: 10.2 micron) at a final Ca/Mg (3:2)concentration of 5 mM. Lowering the ion concentration to 1 mM induces an expansion of the average nuclear diameter to 12.2 micron. Both contracted and expanded nuclei are surrounded by a largely intact nuclear envelope as revealed by thin-sectioning electron microscopy. Nuclear swelling is accompanied by an expansion of the nuclear envelope as indicated by the decrease in the frequency of nuclear pore complexes from 52.6 to 42.1 pores/micron2 determined by freeze-etch electron microscopy. Contracted nuclear membranes reveal particle-devoid areas (average size: 0.21 micron2) on 59% of their fracture faces at the optimal growth temperature of 28 degrees C. About three-fifths of the number of these smooth areas disappear upon nuclear membrane expansion. Electron spin resonance using 5-doxylstearic acid as a spin label indicates a higher lipid fluidity in contracted than in expa,ded nuclear membranes. Moreover, a thermotropic lipid clustering occurs at approximately 17 degrees C only in expanded nuclear membranes. In contrast to the nuclear membrane- bound lipids, free lipids extracted from the nuclei rigidify with increasing Ca/Mg concentrations. Our findings are compatible with the view that the peripheral layer of the fundamental nuclear protein- framework, the so-called nuclear matrix, can modulate, inter alia, the lipid distribution and fluidity, respectively, in nuclear membranes. We suggest that a contraction of the nuclear matrix's peripheral layer induces a contraction of the nuclear membranes which, in turn, leads to an isothermic lateral lipid segregation within nuclear membranes. 相似文献