Role of the capsule produced by Bacillus megaterium ATCC 19213 in the accumulation of metallic cations

A study was undertaken to determine if the capsule produced by Bacillus megaterium ATCC 19213 was capable of binding metallic ions. For non-toxic metallic ions, this was accomplished by determining the relative concentrations of Fe2+, Ca2+, Zn2+, Mg2+, and Mn2+ removed from a chemically defined medium by the normally capsulated parent strain and two mutants with much smaller capsules. For toxic metals, the rates of respiration of the parent strain and a small capsule mutant in the presence of Cu2+, Hg2+, and Ag1+ were compared. It was found that the parent strain accumulated more Ca2+, Mg2+, and Mn2+. Accumulation of Fe2+ and Zn2+ was similar for the parent strain and the small capsule mutants. Respiration of the parent strain was less inhibited by Cu2+, Hg2+, and Ag1+, indicating that these metals are also bound to the capsule.     

Studies on the physical state of water in living cells and model systems. VII. Exclusion of sugars and sugar alcohols from the water in sulfonate ion exchange resins: the "size rule"

The equilibrium distribution of D-arabinose, glycerol, D-ribose, sorbitol, sucrose or inulin in the water of sulfonate ion exchange resins (containing as counterions to the sulfonate anions, H+, Li+, Na+, K+, Rb+, Cs+, NH4+, tetraethylammonium or tetra-n-butyl-ammonium) was studied, usually at one temperature (25 degrees C) but sometimes at two (25 degrees C, 0 degrees C). The apparent equilibrium distribution (p-value) changes with the nature of the counterion, temperature and the molecular weights of the sugar, sugar alcohols, and derivative in question. Linear correlation coefficients between the molecular weights of the solutes and their respective p-values in the range of -0.92 to -0.93 were obtained when the resins were in the Li+, Na+, K+, or Rb+ form.     

TI-205 nuclear magnetic resonance determination of the thermodynamic parameters for the binding of monovalent cations to gramicidins A and C.

Thermodynamic parameters for the binding of the monovalent cations, Li+, Na+, K+, Rb+, Cs+, NH4+, TI+, and Ag+, to gramicidin A and for the binding of TI+ to gramicidin C, incorporated into lysophosphatidylcholine, have been determined using a combination of TI-205 nuclear magnetic resonance spectroscopy and competition binding. The thermodynamic parameters, enthalpy and entropy, are discussed in terms of a process involving the transfer of cations from an aqueous to amide environment.     

Functional role and affinity of inorganic cations in stabilizing the tetrameric structure of the KcsA K+ channel

Crystal structures of the tetrameric KcsA K+ channel reveal seven distinct binding sites for K+ ions within the central pore formed at the fourfold rotational symmetry axis. Coordination of an individual K+ ion by eight protein oxygen atoms within the selectivity filter suggests that ion-subunit bridging by cation-oxygen interactions contributes to structural stability of the tetramer. To test this hypothesis, we examined the effect of inorganic cations on the temperature dependence of the KcsA tetramer as monitored by SDS-PAGE. Inorganic cations known to permeate or strongly block K+ channels (K+, Rb+, Cs+, Tl+, NH4+, Ba2+, and Sr2+) confer tetramer stability at higher temperatures (T0.5 range = 87 degrees C to >99 degrees C) than impermeant cations and weak blockers (Li+, Na+, Tris+, choline+; T0.5 range = 59 degrees C to 77 degrees C). Titration of K+, Ba2+, and other stabilizing cations protects against rapid loss of KcsA tetramer observed in 100 mM choline Cl at 90 degrees C. Tetramer protection titrations of K+, Rb+, Cs+, Tl+, and NH4+ at 85 degrees C or 90 degrees C exhibit apparent Hill coefficients (N) ranging from 1.7 to 3.3 and affinity constants (K0.5) ranging from 1.1 to 9.6 mM. Ba2+ and Sr2+ titrations exhibit apparent one-site behavior (N congruent with 1) with K0.5 values of 210 nM and 11 microM, respectively. At 95 degrees C in the presence of 5 mM K+, titration of Li+ or Na+ destabilizes the tetramer with K0.5 values of 57 mM and 109 mM, respectively. We conclude that specific binding interactions of inorganic cations with the selectivity filter are an important determinant of tetramer stability of KscA.     

Polymetallic Stress Changes the Endogenous Status of Brassinosteroids and Reduces the Effectiveness of Photochemical Reactions Photosystem II in Barley Plants

Doklady Biochemistry and Biophysics - The effect of polymetallic stress (Mn2+, Cd2+, Cu2+, Ni2+, Zn2+, Pb2+, and Al3+) of different duration and intensity on the endogenous brassinosteroids (BS)...     

Effect of metal ions on the stable adduct formation of 16alpha-hydroxyestrone with a primary amine via the Heyns rearrangement.

16alpha-Hydroxyestrone (16alpha-OHE1), one of the major estrogen metabolites in humans that may plays a role in cell transformation, has been found to form stable adducts with nuclear proteins. The mechanism for the formation of a stable covalent adduct of 16alpha-OHE1 with protein has been postulated via the Heyns rearrangement after Schiff base formation. The Heyns rearrangement on the steroidal D-ring alpha-hydroxyimine was investigated using 17-(2-methoxyethylimino)estra-1,3,5(10)-triene-3,16alpha-dio l as a model intermediate. Rates of the Heyns rearrangement and hydrolysis of the steroidal a-hydroxyimine were determined by a high-performance liquid chromatography (HPLC) simultaneously. The Heyns rearrangement was demonstrated to be optimum at pH 6.2 and the reaction rate at physiological pH, 7.3-7.5, was more than 90% of that at the optimum pH. On the other hand, modulator(s) to the reactions were also examined. According to our previous finding of the proton-mediated mechanism of the Heyns rearrangement, the effects of cationic metal ions on the reactions were examined with 29 metal chlorides. Five metal ions, Pt4+, Cu2+, Ni2+, Co2+, and Mn2+, suppressed the formation of Heyns product significantly while Fe2+, Y3+, Gd3+, and Er3+ slightly increased it. The suppression rate was synergistically enhanced by the combination of Pt4+ with Co2+, Cu2+, or Ni2+. These results suggest the five metal ions, Pt4+, Cu2+, Ni2+, Co2+, and Mn2+, reduce the formation of the Heyns product in vivo and, therefore, would be useful tools to clarify the implication of the stable adduct formation of 16alpha-OHE1 with protein.     

A bar model for the pump and channel function of the reconstituted Na+,K+-ATPase

Purified Na+,K+-ATPase is treated with trypsin. The altered enzyme is then reconstituted into liposomes and the change in active and passive Na+,K+-fluxes is recorded. Trypsin treatment transforms the slow passive Na+,K+-fluxes into leaks. The leak formation is correlated with the degree of proteolysis and the associated decrease in Na+,K+-ATPase activity. The active Na+,K+-transport capacity decreases in parallel with the passive transport. It is thus proposed that the Na+,K+-ATPase molecule primarily contains unspecific transmembrane tunnels that are rendered ion-selective by transverse bars of specific length (bar model).     

Ionic composition of endolymph and perilymph in the inner ear of the oyster toadfish, Opsanus tau

The concentrations of free Na+, K+, Ca(+, and Cl(-)in endolymph and perilymph from the inner ear of the oyster toadfish, Opsanus tau, were measured in vivo using double-barreled ion-selective electrodes. Perilymph concentrations were similar to those measured in other species, while endolymph concentrations were similar to those measured previously in elasmobranch fish, though significantly different from concentrations reported in mammals. Perilymph concentrations (mean +/- std. dev.) were as follows: Na+, 129 mmol l(-1) +/- 20; K+, 4.96 mmol l(-1) +/- 2.67; Ca2+, 1.83 mmol l(-1) +/- 0.27; and Cl(-), 171 mmol l(-1) +/- 20. Saccular endolymph concentrations were Na+, 166 mmol l(-1) +/- 22; K+, 51.4 mmol l(-1) +/- 16.7; Ca2+, 2.88 mmol l(-1) +/- 0.27; and Cl(-), 170 mmol l(-1) +/- 12; and semicircular canal (utricular vestibule) endolymph concentrations were Na+, 122 mmol l(-1) +/- 15; K+, 47.7 mmol l(-1) +/- 13.2; Ca2+, 1.78 mmol l(-1) +/- 0.48; Cl(-), 176 mmol l(-1) +/- 27. The relatively high concentrations of Ca2+ and Na+ in the endolymph may have significant implications for the physiological function of the mechanoelectrical transduction channels in the vestibular hair cells of fish compared to those of their mammalian counterparts.     

Cu2+-induced permeability of the Escherichia coli cytoplasmic membrane]

Cu(2+)-induced permeability of cytoplasmic membranes of Escherichia coli for different cations and neutral molecules of saccharose was estimated by studying their effect on cell plasmolysis during uncharged exchange of cytoplasmic K+ ions by periplasmic space cations. The addition of copper resulted in the exchange of K+ ions by periplasmic Na+, Tris+, streptomycin2+, Cu2+, Ca2+, Mg2+, Cd2+, and Mn2+. It is concluded that Cu(2+)-induced conducting pathways in bacterial membranes are hydrophilic channels with a radius of approximately 0.5 nm and a nonselective permeability for different cations.     

Effect of thyroid hormone receptors in normal and cancerous cells on the ionic conductivity of bilayer phospholipid membranes

The effect of thyroid hormones receptors isolated from normal and cancer cells on bilayer phospholipid membranes (BPhLM) conductivity, has been studied. The receptor isolated from normal cells in complex X with the hormone selectively induces H+-conductivity of BPhLM generating transmembrane potential equal to 42 mV on the membrane at pH gradient equal to 1. In the presence of K+, Na+, Ca+, Mn2+, Sr2+, Mg2+ the changes of BPhLM are not observed. Neither hormones (T3, T4) nor receptor in free position affect the BPhLM conductivity. Thyroid hormone receptor isolated from mamalignantly transformed cells in a complex with T3 or T4 increases the BPhLM permeability for Ca2+. The transmembrane potential measured at 10fold Ca2+ ion concentration is equal to 16 mV. In the presence of H+, K+, Na+, Mn2+, Sr2+, Mg2+, Ba2+, the resistance of BPhLM doesn't change.     

Effect of changing temperature on the ionic permeation through the cyclic GMP-gated channel from vertebrate photoreceptors.

Native cGMP-gated channels were studied in rod outer segments of the larval tiger salamander Ambystoma tigrinum. The alpha subunit of the cGMP-gated channel from bovine rods, here referred to as the wild type (w.t.), and mutant channels were heterologously expressed in Xenopus laevis oocytes. These channels were studied in excised membrane patches in the inside-out configuration and were activated by the addition of 100 or 500 microM cGMP. The effect of temperature on the ionic permeation was studied. The macroscopic current flowing through the native channel at +100 mV had an activation energy of 35.8, 30, 31.8, 34.5, 41.3, and 22.4 kJ mol-1 in the presence of Li+, Na+, K+, Rb+, Cs+, and NH4+, respectively. The macroscopic current flowing through the w.t. channel at +100 mV had an activation energy of 45.2, 38.2, 37.5, 47.3, 49.4, and 38.9 kJ mol-1 in the presence of Li+, Na+, K+, Rb+, Cs+, and NH4+, respectively. The activation energy of the macroscopic current flowing through the native and w.t. channels did not vary significantly when the ionic concentration of the permeant ion was changed between 2.5 and 110 mM. The activation energy of the single-channel current of the w.t. channel at +100 mV was 40.4 and 33 kJ mol-1 for Na+ and NH4+, respectively. The reversal potential of biionic solutions changed significantly with temperature. These results can be used to obtain an estimate of the enthalpic and entropic contributions to the barrier of the Gibbs free energy experienced by an ion during its permeation through the open channel. These estimates indicate that the ionic permeation and selectivity of the cGMP-gated channel are controlled both by enthalpic and entropic factors and that the selectivity of the native channel for Li+ over Na+ is primarily caused by entropic effects.     

Registration of changes in the nucleoide conformation based on the analysis of the time dependence of viscosity

The sensitivity of the method of anomalous time dependence of viscosity to changes in the conformation of DNA-protein complexes (such as nucleoide) by the action of (1) nalidixic acid on cells and (2) one- and two-charged ions Na+, Cl-, Cu2+, Ca2+, Mg2+, Zn2+, SO4(2-) on Escherichia coli cells has been studied. The data obtained suggest that the method anomalous time dependence of viscosity is highly sensitive to the conformation of high-molecular DNA-protein complexes. It was shown that the method anomalous time dependence of viscosity can be used to register cell sensitivity to changes in the concentration of one- and two-charged ions in suspension.     

The effect of ions on the adhesion and internalization of Chlamydia trachomatis by HeLa cells

The adhesion and internalization of Chlamydia trachomatis by HeLa cells was unaffected by removal of K+, Mg2+, or glucose from the incubation medium, slightly reduced by removal of Na+, and significantly reduced by omission of Ca2+, Sr2+, Mg2+, and Mn2+ could replace Ca2+ in the adhesion but only Sr2+ supported internalization, and La3+, Co2+, Fe3+, Ba2+, and Zn2+ all reduced internalization more than adhesion. During initial infection there was no measurable difference in the uptake or release of 45Ca2+ or 86Rb+ between infected and noninfected HeLa monolayers. Infection was not prevented by pretreatment of the monolayers with the calcium channel blockers, verapamil, D600, and nitrendipine, or the calmodulin inhibitors, TMB-8 or trifluperazine. The results suggest that divalent cations are not essential for chlamydial infection but that the process of internalization is facilitated by the presence of cations, particularly Na+ and Ca2+.     

Effect of partial hepatectomy on the invertor mechanism of regulation of the Na+,K+-ATPase activity in hepatocytes of rats of various ages]

Age peculiarities of partial hepatectomy effect on the hepatocytes plasma membrane Na+, K(+)-ATPase activity and its insulin-induced stimulation has been studied. It has been shown that partial hepatectomy does not change basal Na+, K(+)-ATPase activity in adult rats. In old partial hepatectomised rats Na+, K(+)-ATPase activity is slightly higher than in control old rats, although this increase is not statistically significant. At the same time, partial hepatectomy acts differently on the insulin-induced Na+, K(+)-ATPase activation in adult and old rats. Insulin activates Na+, K(+)-ATPase at the same extent both in control and partial hepatectomized adult animals. In old hepatectomized rats, but not in old control animals, insulin stimulates Na+, K(+)-ATPase activity as well as. Thus hepatectomy "rejuvenates" old hepatocytes and results in recovery of invertor mechanism of Na+, K(+)-ATPase activation.     

Characteristics of the ionic composition and ultrastructure of Bordetella pertussis in controlled regulation of the mineral element content in a growth medium

The content of trace elements necessary for the normal growth of bacteria was found to have no effect on the intracellular concentration of Ca2+ and Al3+. The content of Cu2+, Fe3+, Mn2+, Mg2+ was considerably reduced. The addition of Mg2+ at different concentrations to this culture medium stimulated the capacity of cells for accumulating not only Mg2+, but also some other ions. Their maximum intracellular concentration was observed when the concentration of Mg2+ in the culture medium was 41 mM. The growth of microbial cells in the standard culture medium containing Mg2+ at a concentration of 4 mM was accompanied by the increased consumption of elements actively participating in redox reactions (Cu2+, Fe3+, Mn2+). Shifts in the ionic composition of microbial cells were manifested by the morphological features of B. pertussis, linked with the increased synthesis of crystalloid structures. The influence of Mn2+, Al3+, Zn2+ at different concentrations on the ionic composition and morphology of B. pertussis was studied.     

外源EBR对NaCl胁迫下燕麦幼苗无机离子吸收、运输和分配的影响

为了探讨外源2,4-表油菜素内酯(2,4-epibrassinolide,EBR)诱导燕麦(Avena sativa L.)幼苗抗盐性的效果及其生理调节机制,以"青引2号"和"加燕2号"燕麦为材料,研究NaCl胁迫下施用外源EBR对燕麦幼苗无机离子吸收、运输和分配的影响。结果表明:100mmol·L-1NaCl胁迫下,"青引2号"和"加燕2号"燕麦幼苗叶片和根系中的Na+、Cl-含量均显著升高,对阳离子的吸收产生了拮抗作用,导致燕麦幼苗叶片和根系中的K+、Ca2+、Mg2+、Mn2+、Fe2+、Zn2+、Cu2+含量显著降低,离子稳态平衡被打破;100 mmol·L-1NaCl胁迫下,施用0.01μmol·L-1外源EBR后,"青引2号"和"加燕2号"燕麦幼苗叶片和根系中的Na+和Cl-含量显著降低,促进了燕麦幼苗根系对K+、Ca2+、Mg2+、Fe2+、Mn2+、Cu2+和Zn2+的吸收,叶片和根系中K+/Na+、Cl-/Na+、Ca2+/Na+、Mg2+/Na+、Fe2+/Na+、Mn2+/Na+、Cu2+/Na+和Zn2+/Na+显著升高,并且有效调控燕麦幼苗体内无机离子的运输比和阳离子的运输选择性比率,离子稳态重新达到平衡状态;说明外源EBR能够缓解NaCl胁迫下Na+和Cl-对燕麦幼苗所造成的离子毒害作用,有效调控燕麦幼苗对无机离子的选择性吸收、运输和分配,对维持燕麦幼苗体内的离子稳态平衡具有正向调控作用。     

Evaluation of the metal-ion-coordinating differences between the 2'-, 3'- and 5'-monophosphates of adenosine

The stability constants of the 1:1 complexes formed between Mg2+, Ca2+, Sr2+, Ba2+, Mn2+, Co2+, Ni2+, Cu2+, Zn2+ or Cd2+ and 2'AMP2-, 3'AMP2- or 5'AMP2- were determined by potentiometric pH titration in aqueous solution (I = 0.1 M, NaNO3; 25 degrees C). The experimental conditions were carefully selected such that self-association of the nucleotides and their complexes is negligibly small; i.e. it was made certain that the properties of the monomeric divalent-metal-ion--AMP [M(AMP)] complexes were studied. Based on recent measurements with simple phosphate monoesters, R-MP2- where R is a non-coordinating residue [Massoud, S. S. & Sigel, H. (1988) Inorg. Chem. 27, 1447-1453], it is shown that all the M(AMP) complexes of the alkaline earth ions, with the possible exception of Mg(5'AMP), have exactly the stability expected for a sole-phosphate coordination of the metal ion. The same property is revealed for the complexes with Mn2+, Co2+, Zn2+ or Cd2+ and 3'AMP2-; in case of Ni(3'AMP) and Cu(3'AMP) a slight stability increase just at the edge of the experimental-error limits is indicated. This slight stability increase is attributed to the formation of a macrochelate (possibly with N-3); in fact, additional information confirms macrochelation for Cu(3'AMP). About 45% of Cu(2'AMP) exists in aqueous solution as a macrochelate (probably involving N-3); the other M(2'AMP) complexes (M2+ = Mn2+, Co2+, Ni2+, Zn2+, Cd2+) form (if at all) only traces of a base-backbound species. Most pronounced is macrochelate formation with 5'AMP2-: all mentioned 3d ions and Zn2+ or Cd2+ form to some extent macrochelates via N-7 (the structures of these closed species are indicated). In case of M(5'AMP) the base-binding site is certain: replacement of N-7 by a CH unit (tubercidin 5'-monophosphate) eliminates any increased complex stability, whereas formation of the 1,N6-etheno bridge to form 1,N6-ethenoadenosine 5'-monophosphate results in the phenanthroline-like N-6,N-7 site which facilitates macrochelation significantly.     

The effect of di- and trivalent metal ions on the binding of [3H-Tyr1, D-Ala2, D-Leu5] enkephalin to opiate receptors from the rat brain

The influence of Ca2+, Mg2+, Mn2+, Sr2+, La3+, Nd3+, Sm3+, Eu3+, and Gd3+ ions on the binding of labeled, stable enkephalin analogue, [3H-Tyr1, D-Ala2, D-Leu5]enkephalin, to opiate receptors of the rat brain membrane preparations has been investigated. The formation of the complex can be described by a scheme involving at least two independent binding sites. The high affinity site does not discriminate the divalent and trivalent metal ions: all examined cations enhanced the enkephalin affinity for this site. The ligand binding to the low affinity site is potentiated only by Mn2+, Mg2+, and lathanoides. The maximal concentration of the binding sites of the above two types is not affected by the cations. The increase in the ionic strength of the solution entails a decrease in the affinity of the ligand for the high affinity binding site. It is shown that the effect of both di- and trivalent metal cations on the [3H-Tyr1, D-Ala2, D-Leu3] enkephalin binding is mediated through one cation attachment site on the respective enkephalin receptor.     

Differences in the protein fluorecence of the two iron(III)-binding sites of ovotransferrin.

1. Changes in the tryptophan fluorescence and the visible absorption spectrum resulting from the combination of apo-ovotransferrin with Fe3+, F,E2+, Cu2+, Zn2+, Mn2+, and Cd2+were measured. 2. As expected for a radiationless transfer of electronic excitation energy, only the ions Fe3+, Fe2+and Cu2+, which gave complexes with large extinctions between 300 and 370nm, resulted in large decreases in trytophan fluorescence. 3. The decrease in protein fluorescence was non-linear with increasing occupancy of the Fe3+ -and Cu2+ - binding sites. The decrease in fluorescence on binding of Fe3+ was biphasic and showed that the two metal-binding sites were being occupied sequentially at pH7.4-8.4. The first site reacted with Fe3+ instantaneously, the second was occupied over a minute. 5. The nonidentity of the two sites was also demonstrated by the preparation of a stable hybrid containing both Cu2+ and Zn2+.h Cu2+ and Zn2+     

Effect of monovalent cations on the catalytic and spectral properties of tyrosine-phenol-lyase from Citrobacter intermedius

The action of monovalent cations Li+, Na+, K+, Rb+, Cs+, NH4+ on catalytic and physico-chemical properties of bacterial tyrosine--phenol-lyase was investigated. It was shown that K+, Rb+, Cs+, NH4+ were the noncompetitive activators of the enzyme, Na+ was an inhibitor, Li+ did not influence the catalytic activity. The values of KA and Vmax were determined for the activators in the reaction of alpha, beta-elimination of L-tyrosine. Monovalent cations affect the absorption and CD spectra of the enzyme and its complex with the quasi-substrate--L-alanine. It was suggested that an activation of tyrosine phenollyase by monovalent cations was connected with the increase of the active protonated form of the holoenzyme (lambda max 420 mm) induced by the cations-activators.