Aggregation of nucleosomes by divalent cations.

Conditions of precipitation of nucleosome core particles (NCP) by divalent cations (Ca(2+) and Mg(2+)) have been explored over a large range of nucleosome and cation concentrations. Precipitation of NCP occurs for a threshold of divalent cation concentration, and redissolution is observed for further addition of salt. The phase diagram looks similar to those obtained with DNA and synthetic polyelectrolytes in the presence of multivalent cations, which supports the idea that NCP/NCP interactions are driven by cation condensation. In the phase separation domain the effective charge of the aggregates was determined by measurements of their electrophoretic mobility. Aggregates formed in the presence of divalent cations (Mg(2+)) remain negatively charged over the whole concentration range. They turn positively charged when aggregation is induced by trivalent (spermidine) or tetravalent (spermine) cations. The higher the valency of the counterions, the more significant is the reversal of the effective charge of the aggregates. The sign of the effective charge has no influence on the aspect of the phase diagram. We discuss the possible reasons for this charge reversal in the light of actual theoretical approaches.     

Effects of Al(3+) and related metals on membrane phase state and hydration: correlation with lipid oxidation

The aim of the present study was to further understand how changes in membrane organization can lead to higher rates of lipid oxidation. We previously demonstrated that Al(3+), Sc(3+), Ga(3+), Be(2+), Y(3+), and La(3+) promote lipid packing and lateral phase separation. Using the probe Laurdan, we evaluated in liposomes if the higher rigidity of the membrane caused by Al(3+) can alter membrane phase state and/or hydration, and the relation of this effect to Al(3+)-stimulated lipid oxidation. In liposomes of dimyristoyl phosphatidylcholine and dimyristoyl phosphatidylserine, Al(3+) (10-100 microM) induced phase coexistence and displacement of T(m). In contrast, in liposomes of brain phosphatidylcholine and brain phosphatidylserine, Al(3+) (10-200 microM) did not affect membrane phase state but increased Laurdan generalized polarization (GP = -0. 04 and 0.09 in the absence and presence of 200 microM Al(3+), respectively). Sc(3+), Ga(3+), Be(2+), Y(3+), and La(3+) also increased GP values, with an effect equivalent to a decrease in membrane temperature between 10 and 20 degrees C. GP values in the presence of the cations were significantly correlated (r(2) = 0.98, P < 0.001) with their capacity to stimulate Fe(2+)-initiated lipid oxidation. Metal-promoted membrane dehydration did not correlate with ability to enhance lipid oxidation, indicating that dehydration of the phospholipid polar headgroup is not a mechanism involved in cation-mediated enhancement of Fe(2+)-initiated lipid oxidation. Results indicate that changes in membrane phospholipid phase state favoring the displacement to gel state can facilitate the propagation of lipid oxidation.     

Kinetic and thermodynamic analysis of the interaction of cations with dialkylglycine decarboxylase

Dialkylglycine decarboxylase (DGD) is a tetrameric pyridoxal phosphate (PLP)-dependent enzyme that catalyzes both decarboxylation and transamination in its normal catalytic cycle. Its activity is dependent on cations. Metal-free DGD and DGD complexes with seven monovalent cations (Li(+), Na(+), K(+), Rb(+), Cs(+), NH(4)(+), and Tl(+)) and three divalent cations (Mg(2+), Ca(2+), and Ba(2+)) have been studied. The catalytic rate constants for cation-bound enzyme (ck(cat) and ck(cat)/bK(AIB)) are cation-size-dependent, K(+) being the monovalent cation with the optimal size for catalytic activity. The divalent alkaline earth cations (Mg(2+), Ca(2+), and Ba(2+)) all give approximately 10-fold lower activity compared to monovalent alkali cations of similar ionic radius. The Michaelis constant for aminoisobutyrate (AIB) binding to DGD-PLP complexes with cations (bK(AIB)) varies with ionic radius. The larger cations (K(+), Rb(+), Cs(+), NH(4)(+), and Tl(+)) give smaller bK(AIB) ( approximately 4 mM), while smaller cations (Li(+), Na(+)) give larger values (approximately 10 mM). Cation size and charge dependence is also found with the dissociation constant for PLP binding to DGD-cation complexes (aK(PLP)). K(+) and Rb(+) possess the optimal ionic radius, giving the lowest values of aK(PLP). The divalent alkaline earth cations give aK(PLP) values approximately 10-fold higher than alkali cations of similar ionic radius. The cation dissociation constant for DGD-PLP-AIB-cation complexes (betaK(M)z+) was determined and also shown to be cation-size-dependent, K(+) and Rb(+) yielding the lowest values. The kinetics of PLP association and dissociation from metal-free DGD and its complexes with cations (Na(+), K(+), and Ba(2+)) were analyzed. All three cations tested increase PLP association and decrease PLP dissociation rate constants. Kinetic studies of cation binding show saturation kinetics for the association reaction. The half-life for association with saturating Rb(+) is approximately 24 s, while the half-life for dissociation of Rb(+) from the DGD-PLP-AIB-Rb(+) complex is approximately 12 min.     

Cd2+ versus Zn2+ uptake by the ZIP8 HCO3--dependent symporter: kinetics, electrogenicity and trafficking

The mouse Slc39a8 gene encodes the ZIP8 transporter, which has been shown to be a divalent cation/HCO3- symporter. Using ZIP8 cRNA-injected Xenopus oocyte cultures, we show herein that: [a] ZIP8-mediated cadmium (Cd(2+)) and zinc (Zn(2+)) uptake have V(max) values of 1.8+/-0.08 and 1.0+/-0.08 pmol/oocyte/h, and K(m) values of 0.48+/-0.08 and 0.26+/-0.09 microM, respectively; [b] ZIP8-mediated Cd(2+) uptake is most inhibited by Zn(2+), second-best inhibited by Cu(2+), Pb(2+) and Hg(2+), and not inhibited by Mn(2+) or Fe(2+); and [c] electrogenicity studies demonstrate an influx of two HCO3- anions per one Cd(2+) (or one Zn(2+)) cation, i.e. electroneutral complexes. Using Madin-Darby canine kidney (MDCK) polarized epithelial cells retrovirally infected with ZIP8 cDNA and tagged with hemagglutinin at the C-terminus, we show that-similar to ZIP4-the ZIP8 eight-transmembrane protein is largely internalized during Zn(2+) homeostasis, but moves predominantly to the cell surface membrane (trafficking) under conditions of Zn(2+) depletion.     

Effect of divalent cations on antiparallel G-quartet structure of d(G4T4G4)

The thermodynamic parameters of an antiparallel G-quartet formation of d(G4T4G4) with 1 mM divalent cation (Mg(2+), Ca(2+), Mn(2+), Co(2+), and Zn(2+)) were obtained. The thermodynamic parameters showed that the divalent cation destabilizes the antiparallel G-quartet of d(G4T4G4) in the following order: Zn(2+)>Co(2+)>Mn(2+)>Mg(2+)>Ca(2+). In addition, a higher concentration of a divalent cation induced a transition from an antiparallel to a parallel G-quartet structure. These results indicate that these divalent cations are a good tool for regulating the G-quartet structures.     

Identification of hyperpolarization-activated calcium channels in apical pollen tubes of Pyrus pyrifolia

The pollen tube has been widely used to study the mechanisms underlying polarized tip growth in plants. A steep tip-to-base gradient of free cytosolic calcium ([Ca(2+)](cyt)) is essential for pollen-tube growth. Local Ca(2+) influx mediated by Ca(2+)-permeable channels plays a key role in maintaining this [Ca(2+)](cyt) gradient. Here, we developed a protocol for successful isolation of spheroplasts from pollen tubes of Pyrus pyrifolia and identified a hyperpolarization-activated cation channel using the patch-clamp technique. We showed that the cation channel conductance displayed a strong selectivity for divalent cations, with a relative permeability sequence of barium (Ba(2+)) approximately Ca(2+) > magnesium (Mg(2+)) > strontium (Sr(2+)) > manganese (Mn(2+)). This channel conductance was selective for Ca(2+) over chlorine (Cl(-)) (relative permeability P(Ca)/P(Cl) = 14 in 10 mm extracellular Ca(2+)). We also showed that the channel was inhibited by the Ca(2+) channel blockers lanthanum (La(3+)) and gadolinium (Gd(3+)). Furthermore, channel activity depended on extracellular pH and pollen viability. We propose that the Ca(2+)-permeable channel is likely to play a role in mediating Ca(2+) influx into the growing pollen tubes to maintain the [Ca(2+)](cyt) gradient.     

Kinetics of ion translocation across charged membranes mediated by a two-site transport mechanism. Effects of polyvalent cations upon rubidium uptake into yeast cells.

(1) The effect of surface charge upon the kinetics of monovalent cation translocation via a two-site mechanism is investigated theroretically. (2) According to the model dealt with, typical relations are expected for the dependence of the kinetic parameters of the translocation process upon the concentration of a polyvalent cation, differing essentially from those derived for the case in which the membrane carries no excess charge. (3) Even when a polyvalent cation does not compete with the substrate cation for binding to the translocation sites, apparently competitive inhibition may occur when the membrane is negatively charged. (4) The model is tested experimentally by studying the effects of the polyvalent cations Mg2+, Sr2+, Ca2+, Ba2+ and Al3+ upon Rb+ uptake into yeast cells at pH 4.5 A good applicability is found. (5) Equimolar concentrations of polyvalent cations reduce the rate of the Rb+ uptake into yeast cells in the order Mg2+ less than Sr2+ less than Ca2+ less than Ba2+ less than Al3+. (6) The conclusion is reached that the reduction in the rate of Rb+ uptake caused by the polyvalent cations applied results mainly from screening of the negative fixed charges on the membrane surface and binding to these negative sites rather than competition with Rb+ for the transport sites. (7) The results of our investigation indicate the affinity of the alkaline-earth cations for the negative fixed charges on the surface to the yeast cell membrane increases in the orther Mg2+ less than Sr2 less than Ca2+ less than Ba2+. (8) Probably mainly phosphoryl groups determine the net charge on the membrane of the yeast cell at a medium pH of 4.5.     

Dipole potentials indicate restructuring of the membrane interface induced by gadolinium and beryllium ions

The dipole component of the membrane boundary potential, phi(d), is an integral parameter that may report on the conformational state of the lipid headgroups and their hydration. In this work, we describe an experimental approach to measurements of the dipole potential changes, Deltaphi(d), and apply it in studies of Be(2+) and Gd(3+) interactions with membranes composed of phosphatidylserine (PS), phosphatidylcholine (PC), and their mixtures. Deltaphi(d) is determined as the difference between the changes of the total boundary potential, phi(b), measured by the IFC method in planar lipid membranes and the surface potential, phi(s), determined from the electrophoretic mobility of liposomes. The Gouy-Chapman-Stern formalism, combined with the condition of mass balance, well describes the ion equilibria for these high-affinity cations. For the adsorption of Be(2+) and Gd(3+) to PC membranes, and of Mg(2+) to PS membranes, the values of Deltaphi(b) and Deltaphi(s) are the same, indicative of no change of phi(d). Binding of Gd(3+) to PS-containing membranes induces changes of phi(d) of opposite signs depending on the density of ionized PS headgroups in the bilayer. At low density, the induced Deltaphi(d) is negative (-30 mV), consistent with the effect of dehydration of the surface. At maximal density (pure PS, neutral pH), adsorption of Be(2+) or Gd(3+) induces an increase of phi(d) of 35 or 140 mV, respectively. The onset of the strong positive dipole effect on PS membranes with Gd(3+) is observed near the zero charge point and correlates with a six-fold increase of membrane tension. The observed phenomena may reflect concerted reorientation of dipole moments of PS headgroups as a result of ion adsorption and lipid condensation. Their possible implications to in-vivo effects of these high-affinity ions are discussed.     

Cations residing at the selectivity filter of the voltage-gated Ca2+-channel modify fusion-pore kinetics

Strontium (Sr(2+)), Barium (Ba(2+)) and Lanthanum (La(3+)) can substitute for Ca(2+) in driving synaptic transmission during membrane depolarization. Ion recognition at the polyglutamate motif (EEEE), comprising the channel selectivity-filter, during voltage-driven transitions, controls the kinetics of the voltage-gated calcium channel (VGCC) and its interactions with the synaptic proteins. We tested the effect of different charge carriers on evoked-release, as a means of exploring the involvement of VGCC in the fusion pore configuration. Employing amperometry recordings in single bovine chromaffin cells we show that the size of the fusion pore, designated by the 'foot'-amplitude, was increased when Ba(2+) substituted for Ca(2+) and decreased, with La(3+). The fusion pore stability, indicated by 'foot'-width, was decreased in La(3+). Also, the mean open time of the fusion pore (tau(fp)) was significantly lower in Sr(2+) and La(3+) compared to Ba(2+) and Ca(2+). These cations when occupying the selectivity filter reduced the spike frequency in the order of Ca(2+) > Sr(2+) > Ba(2+) > La(3+), which is parallel to the reduction in total catecholamine release. The correlation between ion binding at the selectivity filter and fusion pore properties supports a model in which the Ca(2+) channel regulates secretion through a site at the selectivity filter, upstream to cation entry into the cell.     

Calcium Requirement for Indoleacetic Acid-induced Acidification by Avena Coleoptiles

The ionic specificity of IAA-induced acidification by Avena coleoptiles was studied, using zwitterionic, presumably impermeant buffers. The acidification was almost totally dependent on divalent cations with an order of effectiveness of Ca(2+) >/= Sr(2+) > Mn(2+), Mg(2+); whereas other polyvalent cations tested were ineffective. The Ca(2+) response was IAA-dependent. The CaCl(2) concentration was optimal at 0.3 to 1 mm and inhibitory at higher concentrations. Sr(2+) inhibited Ca(2+)-dependent acidification and monovalent cations such as K(+) did not induce additional acidification in the presence of optimal CaCl(2). These data are consistent with a mechanism for IAA-induced acidification involving a Ca(2+) -H(+) exchange.     

Sensing of extracellular calcium by neurones

Transient changes in the intracellular concentration of Ca2+ provide a major signal for the regulation of many ion channels and enzymes in central neurones. In contrast, changes in extracellular Ca2+ are thought to play little or no signaling role. However, concentrations of extracellular calcium in the central nervous system do change dramatically during intense physiological and pathological stimulation, and recent studies have identified a number of membrane proteins that can sense and respond to changes in extracellular Ca2+. These include the recently cloned Ca(2+)-sensing receptor, hemi-gap-junction channels, and a potential Ca(2+)-sensing cation channel. Lowering extracellular Ca2+ strongly depolarizes and excites cultured hippocampal neurones. The excitation can be detected with decreases from physiological concentrations of as little as 100 microM. The depolarization results from activation of a nonselective cation current, which is sensitive to block by divalent and polyvalent cations. In outside-out patches, lowering Ca2+ induces a single-channel current with a conductance of 36 pS. Activation of this cation channel, in response to decreases in extracellular Ca2+, likely plays a key role in a positive feedback system of excessive neuronal depolarization, which accompanies intense excitatory activity in the hippocampus.     

Mannose-Escherichia coli interaction in the presence of metal cations studied in vitro by colorimetric polydiacetylene/glycolipid liposomes

Supramolecular assemblies of liposomes (vesicles) made of diacetylenic lipids and synthetic mannoside derivative glycolipid receptors were successfully used to mimic the molecular recognition occurring between mannose and Escherichia coli. This specific molecular recognition was translated into visible blue-to-red color transition (biochromism) of the polymerized liposomes, readily quantified by UV-visible spectroscopy. Some transition metal cations (Cd(2+), Ag(+), Cu(2+), Fe(3+), Zn(2+) and Ni(2+)) and alkali earth metal cations (Ca(2+), Mg(2+) and Ba(2+)) were introduced into the system to analyze their effects on specific biochromism. Results showed that the presence of Cd(2+), Ag(+), Ca(2+), Mg(2+) and Ba(2+) enhanced biochromism. A possible enhancement mechanism was proposed in the process of bacterial adhesion to host cells. However, Cu(2+), Fe(3+), Zn(2+) and Ni(2+) exhibited inhibitory effects that cooperated with diacetylene lipid with a carboxylic group and increased the rigidity of the liposomal outer leaflet, blocking changes in the side chain conformation and electrical structure of polydiacetylene polymer during biochromism.     

Interaction of the alternating double stranded copolymer poly(dA-dT) x poly(dA-dT) with NiCl(2) and CdCl(2): solution behavior

The thermal denaturation of the synthetic high molecular weight double stranded polynucleotide poly(dA-dT) x poly(dA-dT) has been studied in aqueous buffered solution (Tris 1.0 mM; pH 7.8+/-0.2) in the presence of increasing concentrations of either Ni(2+) (borderline cation) or Cd(2+) (soft cation) at four different constant ionic strength values (NaCl), making use of UV and circular dichroism (CD) spectroscopies. The experimental results show that the B-type double helix of the polymer is stabilized against thermal denaturation in the presence of both cations at low concentrations, relative to the systems where only NaCl is present, in the same conditions of ionic strength and pH. The effect is more pronounced for Ni(2+) than for Cd(2+). At higher concentrations, both cations start to destabilize the double helix, with Cd cations inducing larger variations of T(m). In many cases, when denaturation starts, interstrand cross-linking occurs with formation of aggregates that precipitate.     

Binding of bivalent cations by xanthan in aqueous solution

The interaction between xanthan and selected bivalent cations (Ca(2+), Mg(2+), Mn(2+), Fe(2+), Cu(2+), Zn(2+), Cd(2+) and Pb(2+)) was studied by means of conductometry, viscometry, and nuclear magnetic resonance spectroscopy. Xanthan from Xanthomonas campestris was studied in comparison with dextran from Leuconostoc mesenteroides. While dextran does not develop specific interactions with the bivalent cations, the analysis of the experimental data shows that xanthan chains (M(n) approximately 1.4x10(5) to 2.9x10(6)g/mol) reversibly bind Me(2+) species in aqueous solution at pH 6. Conductometric and viscometric titrations show that a single bivalent cation forms a complex which involves two disaccharide units of the main chain together with two side chains. Based on dipolar magnetic interactions between Mn(2+) and individual carbon positions of xanthan, a possible structure of a chelate-like complex is proposed which involves the pyruvate units at the terminal ends of the side chains as the main binding sites. According to the stoichiometric relation between cations and disaccharide units, a single bivalent cation is bound between the terminal ends of two side chains, leading to an intramolecular cross-link and a reduced hydrodynamic radius of the overall macromolecule. The results indicate that heavy metal ions (Cd(2+) and Pb(2+)) link stronger to the xanthan chain than lighter cations (Ca(2+) and Mg(2+)), a fact which may be of ecological relevance.     

Dual actions of lanthanides on ACTH-inhibited leak K(+) channels

Bovine adrenal zona fasciculata cells express background K(+) channels (I(AC) channels) whose activity is potently inhibited by ACTH. In whole cell patch clamp recordings, it was discovered that the trivalent lanthanides (Ln(3+)s) lanthanum and ytterbium interact with two binding sites to modulate K(+) flow through these channels. Despite large differences in ionic radii, these Ln(3+)s inhibited I(AC) channels half-maximally with IC(50) values near 50 microM. In addition, these Ln(3+)s blocked and reversed ACTH-mediated inhibition of I(AC) K(+) channels at similar concentrations. The Ln(3+)s did not alter inhibition of I(AC) by angiotensin II or cAMP. Ln(3+)-induced uncoupling of ACTH receptor activation from I(AC) inhibition was prevented by raising the external Ca(2+) concentration from 2 to 10 mM. The divalent cation Ni(2+) (500 microM) also blocked ACTH-dependent inhibition of I(AC) through a Ca(2+)-sensitive mechanism. The results are consistent with a model in which Ln(3+)s produce opposing actions on I(AC) K(+) currents through two separate binding sites. In addition to directly inhibiting I(AC), Ln(3+)s (and Ni(2+)) bind with high affinity to a Ca(2+)-selective site associated with the ACTH receptor. By displacing Ca(2+) from this site, Ln(3+)s prevent ACTH from binding and accelerate its dissociation. These results identify Ln(3+)s as a relatively potent group of noncompetitive ACTH receptor antagonists. Allosteric actions of trivalent and divalent metal cations on hormone binding, mediated through Ca(2+)-specific sites, may be common to a variety of peptide hormone receptors.     

A cation channel for K+ and Ca2+ from Tetrahymena cilia in planar lipid bilayers

The single-channel properties for monovalent and divalent cations of a voltage-independent cation channel from Tetrahymena cilia were studied in planar lipid bilayers. The single-channel conductance reached a maximum value as the K+ concentration was increased in symmetrical solutions of K+. The concentration dependence of the conductance was approximated to a simple saturation curve (a single-ion channel model) with an apparent Michaelis constant of 16.3 mM and a maximum conductance of 354 pS. Divalent cations (Ca2+, Ba2+, Sr2+, and Mg2+) also permeated this channel. The sequence of permeability determined by zero current potentials at high ionic concentrations was Ba2+ greater than or equal to K+ greater than or equal to Sr2+ greater than Mg2+ greater than Ca2+. Single-channel conductances for Ca2+ were nearly constant (13.9 pS-20.5 pS) in the concentrations between 0.5 mM and 50 mM Ca-gluconate. In the experiments with mixed solutions of K+ and Ca2+, a maximum conductance of Ca2+ (gamma Camax) and an apparent Michaelis constant of Ca2+ (K Cam) were obtained by assuming a simple competitive relation between the cations. Gamma Camax and K Cam were 14.0 pS and 0.160 mM, respectively. Single-channel conductances in mixed solutions were well-fitted to this competitive model supporting that this cation channel behaves as a single-ion channel. This channel had relatively high-affinity Ca2+-binding sites.     

Calorimetric study of the interaction of the C2 domains of classical protein kinase C isoenzymes with Ca2+ and phospholipids

The affinities of Ca(2+) and anionic lipid vesicles from the C2 domains of classical protein kinase C subfamily (alpha, betaII, and gamma) were studied using isothermal titration calorimetry (ITC). In addition, the thermal stability of these C2 domains in the presence of different ligand concentrations was analyzed using differential scanning calorimetry (DSC). These three closely related C2 domains bind Ca(2+) in a similar way, demonstrating the presence of two sets of sites. The first set of sites binds one Ca(2+) ion exothermically with similar high affinity for the three proteins (K(d) around 1 microM), while the second set of sites binds endothermically approximately two Ca(2+) ions with lower affinity, which varies for each C2 domain: 22.2 microM for the PKCalpha-C2 domain, 17.2 microM for the PKCbetaII-C2 domain, and 4.3 microM for the PKCgamma-C2 domain. In the absence of Ca(2+), the three C2 domains showed a weak interaction with vesicles containing anionic phospholipids. However, in the presence of a saturating Ca(2+) concentration, the C2 domains increased their affinities for the anionic lipid vesicles. In all cases, the C2 domains bound the vesicles exothermically and with similar affinities. A DSC thermal stability study of the C2 domains in the presence of Ca(2+) and anionic lipids provided further information about this protein-ligand interaction. The presence of increasing Ca(2+) concentrations was matched by an increase in the T(m) in all cases, which was even greater in the presence of anionic lipid vesicles. The extent of the change in T(m) differed for each C2 domain, reflecting the differing effect of the ligands bound during the protein stabilization. Denaturation of the C2 domains was irreversible both in the absence and in the presence of ligands, although the thermograms were not kinetically controlled. The dependence of the T(m) on the Ca(2+) concentration indicates that the protein stabilization observed by DSC primarily reflects the saturation by the cation of the low-affinity set of sites.     

Understanding the multiple functions of Nramp1

Nramp1 regulates macrophage activation in infectious and autoimmune diseases. Nramp2 controls anaemia. Both are divalent cation (Fe(2+), Zn(2+), and Mn(2+)) transporters; Nramp2 a symporter of H(+) and metal ions, Nramp1 a H(+)/divalent cation antiporter. This provides a model for metal ion homeostasis in macrophages. Nramp2, localised to early endosomes, delivers extracellularly acquired divalent cations into the cytosol. Nramp1, localised to late endosomes/lysosomes, delivers divalent cations from the cytosol to phagolysosomes. Here, Fe(2+) generates antimicrobial hydroxyl radicals via the Fenton reaction. Zn(2+) and Mn(2+) may also influence endosomal metalloprotease activity and phagolysosome fusion. The many cellular functions dependent on metal ions as cofactors may explain the multiple pleiotropic effects of Nramp1, and its complex roles in infectious and autoimmune disease.     

Oxidative modification of rat liver 5'-nucleotidase: the mechanisms for protection and re-activation

The effect of oxidative stress catalysed by transition metals appears to have a critical relevance for the structure and function not only of membrane lipids but also of integral membrane proteins in a complex lipid-protein assembling, and membrane-dependent function. The integral membrane enzyme 5'-nucleotidase is susceptible to Fe((2+))-ion catalysed oxidative modification, and the extent of enzyme inhibition is in inverse relationship (r = -0.820) with lipid peroxidation (MDA) level. This work is also a comparative study about possible effectiveness of different Fe-ion chelators (deferoxamine, Na-citrate, Na-salicylate, ammonium oxalate and EDTA), antioxidants (GSH, GSH/GSH-Px system, Cu, Zn-SOD and mannitol) and metal cations (Mg(2+) and Mn(2+)) to protect or restore Fe(2+)-ion induced 5'-nucleotidase inhibition and to suppress Fe(2+)-ion enhanced lipid peroxidation. Among the examined chelators it was only deferoxamine and Na-citrate that exerted a fully protective and reactivating ability; among the antioxidants it was only GSH; among the metal cations it was only Mn(2+). The ability to protect or restore 5'-nucleotidase activity and to diminish chain-induced lipid peroxidation is explicable in terms of: metal-binding ability, capacity of taking iron away from a biological molecule, or ability of transferring the damage to itself. After a short incubation period, the iron associated with enzyme or lipid hydroperoxides could be in a labile coordinative linkage, still able to interact with possible ligands or metal cations.     

The formation of non-bilayer structures in total polar lipid extracts of chloroplast membranes

The structural organisation of aqueous dispersions of total membrane lipid extracts of broad bean (Vicia faba) chloroplasts is dependent on pH and the presence of cations. In the absence of inorganic salts, sonicated dispersions of lipid extract in distilled water form smooth, single-shell vesicles approximately 30–50 nm in diameter. Reducing the pH of the dispersions, to neutralise the acidic lipids present in the extract, or the addition of low concentrations of metal cations, leads to the fusion of the vesicles and a partial phase-separation of the non-bilayer forming lipid monogalactosyldiacylglycerol to form spherical inverted micelles similar to those previously reported for binary mixtures of monogalactosyl and digalactosyldiacylglycerol (Biochim. Biophys. Acta 685, 297–306). Increasing concentrations of polyvalent, but not monovalent, cations lead to further structural rearrangements involving the formation of para-crystalline arrays of tubular and spherical inverted micelles. The factors determining the formation of these different structures, and their possible relevance to the structural organisation of the native chloroplast membrane, are discussed.