Membrane affinity of the amphiphilic marinobactin siderophores

Marinobactins are a class of newly discovered marine bacterial siderophores with a unique amphiphilic structure, suggesting that their functions relate to interactions with cell membranes. Here we use small and large unilamellar L-alpha-dimyristoylphosphatidylcholine vesicles (SUVs and LUVs) as model membranes to examine the thermodynamics and kinetics of the membrane binding of marinobactins, particularly marinobactin E (apo-M(E)) and its iron(III) complex, Fe-M(E). Siderophore-membrane interactions are characterized by NMR line broadening, stopped-flow spectrophotometry, fluorescence quenching, and ultracentrifugation. It is determined that apo-M(E) has a strong affinity for lipid membranes with molar fraction partition coefficients K(x)()(apo)(-)(M)E = 6.3 x 10(5) for SUVs and 3.6 x 10(5) for LUVs. This membrane association is shown to cause only a 2-fold decrease in the rate of iron(III) binding by apo-M(E). However, upon the formation of the iron(III) complex Fe-M(E), the membrane affinity of the siderophore decreased substantially (K(x)()(Fe)(-)(M)E = 1.3 x 10(4) for SUVs and 9.6 x 10(3) for LUVs). The kinetics of membrane binding and dissociation by Fe-M(E) were also determined (k(on)(Fe)(-)(M)E = 1.01 M(-)(1) s(-)(1); k(off)(Fe)(-)(M)E = 4.4 x 10(-)(3) s(-)(1)). The suite of marinobactins with different fatty acid chain lengths and degrees of chain unsaturation showed a range of membrane affinities (5.8 x 10(3) to 36 M(-)(1)). The affinity that marinobactins exhibit for membranes and the changes observed upon iron binding could provide unique biological advantages in a receptor-assisted iron acquisition process in which loss of the iron-free siderophore by diffusion is limited by the strong association with the lipid phase.     

Synthesis, structure, and molecular dynamics of gallium complexes of schizokinen and the amphiphilic siderophore acinetoferrin

A new general synthesis of the citrate-based siderophores acinetoferrin (Af) and schizokinen (Sz) and their analogues is described. The molecular structure of gallium schizokinen, GaSz, was determined by combined (1)H NMR, Hartree-Fock ab initio calculations, DFT, and empirical modeling of vicinal proton NMR spin-spin couplings. The metal-coordination geometry of GaSz was determined from NOE contacts to be cis-cis with respect to the two chelating hydroxamates. One diaminopropane adopts a single chairlike conformation while another is a mixture of two ring pucker arrangements. Both amide hydrogens are internally hydrogen bonded to metal-ligating oxygen atoms. The acyl methyl groups are directed away from each other with an average planar angle of ca. 130 degrees. The kinetics of GaSz racemization were followed by selective, double spin-echo inversion-recovery (1)H NMR spectroscopy over the temperature range of 10-45 degrees C. The racemization proceeds by a multistep mechanism that is proton independent between pD 5 and 12 (k(0) = 1.47 (0.15 s(-1))) and acid catalyzed below pD 4 (k(1) = 2.25 (0.15) x 10(4) M(-1) s(-1)). The activation parameters found for the two sequential steps of the proton independent pathway were DeltaH(++) = 25 +/- 3 kcal M(-1), DeltaS(++) = 25 +/- 7 cal M(-1) K(-1) and DeltaH(++) = 17.1 +/- 0.2 kcal M(-1), DeltaS(++) = 0.3 +/- 2.7 cal M(-1) K(-1). The first step of the proton-independent mechanism was assigned to the dissociation of the carboxyl group. The second step was assigned to complex racemization. The proton-assisted step was assigned to a complete dissociation of the alpha-hydroxy carboxyl group at pD < 4. The ab initio modeling of gallium acinetoferrin, GaAf, and analogues derived from the structure of GaSz has shown that the pendant trans-octenoyl fragments are oriented in opposite directions with the average planar angle of ca. 130 degrees. This arrangement prevents GaAf from adopting a phospholipid-like structural motif. Significantly, iron siderophore complex FeAf was found to be disruptive to phospholipid vesicles and is considerably more hydrophilic than Af, with an eight-fold smaller partition coefficient.     

Interaction of 2-chloronaphthalene with high carbon iron filings (HCIF): adsorption, dehalogenation and mass transfer limitations

Interaction of 2-chloronaphthalene (2-CN) with high-carbon iron filings (HCIF) was studied in anaerobic batch systems, both under well-mixed and poorly-mixed conditions. In well-mixed conditions, partitioning of 2-CN between solid and aqueous phases was fast, resulting in rapid attainment of equilibrium. Equilibrium partitioning could be described by a Freundlich isotherm, C(s)=K x [C(a)](m), where C(s) (micromoles g(-1) iron) and C(a) (micromoles L(-1)) were the solid and aqueous phase 2-CN concentrations, respectively. Isotherm parameters, m and K were determined to be 0.76 and 5.6 x 10(-2) (micromole g(-1) iron)/(micromole L(-1)), respectively. Sorption (k(2)) and desorption (k(3)) rate constants were determined to be 5.60 x 10(-1) h(-1) g(-1) iron L and 10 h(-1), respectively. Reductive dehalogenation of aqueous phase 2-CN occurred concurrently but at a slower rate, and could be described by the expression (dC(T)//dt)= -k(1) x M x (C(a))(N), where C(T) (micromoles L(-1)) was the total 2-CN concentration and M (g iron L(-1)) the concentration of HCIF. The values of k(1) and N were determined to be 1.09 x 10(-2) h(-1) g(-1) iron L and 1.647, respectively. In poorly mixed conditions, adsorption (k(2)) and desorption (k(3)) rate constants were 3.92 x 10(-5) h(-1) g(-1) iron L and 7 x 10(-4) h(-1), respectively, i.e., several orders of magnitude less than in well-mixed systems. The dehalogenation rate parameters, k(1) and N were determined to be 2.22 x 10(-4) h(-1) g(-1) iron L and 0.986, respectively, suggesting slower dehalogenation. These results highlight how mass-transfer limitations during the interaction between HCIF and 2-CN in poorly mixed systems, such as permeable reactive barriers (PRBs), can potentially impact the dehalogenation process.     

Aggregation and fusion of phospholipid vesicles

Membrane fusion and aggregation of phospholipid vesicles are reviewed and discussed. The fusion process is viewed as consisting of several stages: aggregation and close apposition of the particles, destabilization, and finally, merging of the bilayers. A procedure is presented which yields both the rate constant of the dimerization (C₁₁) and the rate constant for fusion of the dimers (f₁₁), which is a direct measure of the probability that two apposed vesicles will fuse. Experimental methods used in the study of membrane fusion are reviewed, primarily with respect to their capacity to monitor the kinetics of vesicle fusion. A few kinetic studies on the mixing of aqueous contents, leakage and increase in size of fusing vesicles are presented in detail.The range of C₁₁ values for Ca²⁺-induced aggregation and fusion of small unilamellar vesicles (SUV, ～ 125 Å radius) composed of phosphatidylserine (PS) is 10⁶ to 5 × 10⁷ M^-1 in the presence of Ca²⁺ concentrations from 1.15 to 2 mM, respectively. For larger PS vesicles (LUV, ～ 500 Å radius) C₁₁ = 6.5 × 10⁷ M^-1s^-1 in the presence of 5 mM Ca²⁺. These values are in good agreement with theoretical calculations based on van der Waals and electrostatic interactions, in which binding of cations is explicitly taken into account. The rate constants of fusion, f₁₁, are 5 s^-1 for PS SUV and 0.08 s^-1 for LUV in the presence of 2 mM and 5 mM Ca²⁺, respectively. The significance of these fusion rate constants to the duration of the fusion event is discussed.Factors affecting fusion such as cations, temperature, membrane composition vesicle concentration and size are reviewed and analyzed. Di- or tri-valent cations induce fusion of acidic phospholipid vesicles (except for phosphatidylinositol) in either pure or mixed form. Among the neutral phospholipids, phosphatidylcholine (PC) inhibits but phosphatidylethanolamine (PE) sustains or enhances the fusion capacity of acidic phospholipid vesicles. Monovalent cations induce reversible aggregation of negatively charged vesicles, but they inhibit the fusion induced by divalent cations such as Ca²⁺ or Mg²⁺. Fusion of neutral phospholipid vesicles, and it occurs the cation-induced fusion of acidic phospholipid vesicles, and it occurs only at temperatures below the gel to liquid crystalline phase transition temperature Tc. This is in contrast to the acidic phospholipid vesicle fusion which is greatly enhanced when the temperature is above the Tc of the phospholipid.     

Synthesis, solution behavior, thermal stability, and biological activity of an Fe(III) complex of an artificial siderophore with intramolecular hydrogen bonding networks

Previously, an artificial siderophore complex, the iron(III) complex with tris[2-[(N-acetyl-N-hydroxy)glycylamino]ethyl]amine (TAGE), was constructed in order to understand the effect of intramolecular hydrogen bonding interaction on the siderophore function, and its structural characterization in the solid state was reported (Inorg. Chem. 2001, 40, 190). In this paper, the solution behavior of the M(III)-TAGE (M = Fe, Ga) system has been investigated using (1)H NMR, UV-vis, and FAB mass spectroscopies in efforts to characterize the biological implication of hydrogen bonding networks between the amide hydrogens and coordinating aminohydroxy oxygens of the complex. The temperature dependence of (1)H NMR spectra for Ga(III) complex of TAGE indicates that hydrogen bonding networks are maintained in polar solvents such as DMSO-d(6) and D(2)O. The UV-vis spectra of the Fe(III)-TAGE system under various pH conditions have shown that TAGE forms a tris(hydroxamato)iron(III) complex in an aqueous solution in the pH range 4-8. By contrast, tris[2-[(N-acetyl-N-hydroxy)propylamido]ethyl]amine (TAPE; TAGE analogue that is difficult to form intramolecular hydrogen bonding networks), which has been synthesized as a comparison of TAGE, forms both of bis- and tris(hydroxamato)iron(III) complexes in the same pH range. Both the stability constants (log beta(FeTAGE) = 28.6; beta(FeTAGE) = [Fe(III)TAGE]/([Fe(3+)][TAGE(3)(-)])) and pM (-log[Fe(3+)]) value for Fe(III)TAGE (pM 25) are comparable to those of a natural siderophore ferrichrome (log beta = 29.1 and pM 25.2). The kinetic study of the TAGE-Fe(III) system has given the following rate constants: the rate of the ligand exchange reaction between Fe(III)TAGE and EDTA is 6.7 x 10(-4) s(-1), and the removal rates of iron from diferric bovine plasma transferrin by TAGE are 2.8 x 10(-2) and 6.0 x 10(-3) min(-1). These values are also comparable to those of a natural siderophore desferrioxamine B under the same conditions. In a biological activity experiment, TAGE has promoted the growth of the siderophore-auxotroph Gram-positive bacterium Microbacterium flavescens, suggesting that TAGE mimics the activity of ferrichrome. These results indicate that the artificial siderophore with intramolecular hydrogen bonding networks, TAGE, is a good structural and functional model for a natural ferrichrome.     

Mechanistic studies of nitric oxide reactions with water soluble iron(II), cobalt(II), and iron(III) porphyrin complexes in aqueous solutions: implications for biological activity.

The reactions of nitric oxide and carbon monoxide with water soluble iron and cobalt porphyrin complexes were investigated over the temperature range 298-318 K and the hydrostatic pressure range 0.1-250 MPa [porphyrin ligands: TPPS = tetra-meso-(4-sulfonatophenyl)porphinate and TMPS = tetra-meso-(sulfonatomesityl)porphinate]. Large and positive DeltaS(double dagger) and DeltaV(double dagger) values were observed for NO binding to and release from iron(III) complexes Fe(III)(TPPS) and Fe(III)(TMPS) consistent with a dissociative ligand exchange mechanism where the lability of coordinated water dominates the reactivity with NO. Small positive values for Delta and Delta for the fast reactions of NO with the iron(II) and cobalt(II) analogues (k(on) = 1.5 x 10(9) and 1.9 x 10(9) M(-1) s(-1) for Fe(II)(TPPS) and Co(II)(TPPS), respectively) indicate a mechanism dominated by diffusion processes in these cases. However, reaction of CO with the Fe(II) complexes (k(on) = 3.6 x 10(7) M(-1) s(-1) for Fe(II)(TPPS)) displays negative Delta and Delta values, consistent with a mechanism dominated by activation rather than diffusion terms. Measurements of NO dissociation rates from Fe(II)(TPPS)(NO) and Co(II)(TPPS)(NO) by trapping free NO gave k(off) values of 6.3 x 10(-4) s(-1) and 1.5 x 10(-4) s(-1). The respective M(II)(TPPS)(NO) formation constants calculated from k(on)/k(off) ratios were 2.4 x 10(12) and 1.3 x 10(13) M(-1), many orders of magnitude larger than that (1.1 x 10(3) M(-1)) for the reaction of Fe(III)(TPPS) with NO.     

Effect of sphingomyelinase-mediated generation of ceramide on aggregation of low-density lipoprotein

This study addresses the response-to-retention hypothesis, which states that the subendothelial retention of atherogenic lipoproteins is the necessary and sufficient condition for the initiation of atherosclerosis. Here we focus on the relationship between the generation of ceramide in the low-density lipoprotein (LDL) phospholipid monolayer and the resulting aggregation of LDL particles. This study provides the first measurement of neutral, Mg (2+)-dependent Sphingomyelinase (Smase)-mediated ceramide formation from LDL-sphingomyelin and does so for a range of enzyme concentrations (0-0.22 units Smase/mL). The kinetics of ceramide generation was measured using a fluorescence assay for the above enzyme concentrations with a fixed substrate concentration (0.33 mg LDL/mL). The kinetics of LDL aggregate formation was measured by dynamic light scattering (DLS, method of cumulants) for identical enzyme concentrations. Ceramide concentration profiles were fit with a modification of the Michaelis-Menten model ( k a = 1.11 x 10 (-1) microM (-1) min (-1), k -a = 6.54 x 10 (2) microM (-1) min (-1), k 1 = 3.33 x 10 (1) microM (-1) min (-1), k -1 = 1.41 x 10 (-2) min (-1), k cat = 8.05 x 10 (1) min (-1), K M = 2.418 microM, k deact = 4.66 x 10 (-2) microM (-1) min (-1)) that accounts for the effects of enzyme attachment to the LDL monolayer and for deactivation of Smase due to product inhibition. LDL aggregation is described by a mass action model as explained in previous studies. A key result of this work is the finding that LDL aggregate size depends directly on ceramide concentration and is independent of enzyme concentration. This study demonstrates how principles of colloid science are relevant to important biomedical problems.     

Thermodynamics, kinetics, and mechanism of the stepwise dissociation and formation of Tris(L-lysinehydroxamato)iron(III) in aqueous acid

pK(a) values for the hydroxamic acid, alpha-NH(3)(+), and epsilon-NH(3)(+) groups of L-lysinehydroxamic acid (LyHA, H(3)L(2+)) were found to be 6.87, 8.89, and 10.76, respectively, in aqueous solution (I = 0.1 M, NaClO(4)) at 25 degrees C. O,O coordination to Fe(III) by LyHA is supported by H(+) stoichiometry, UV-vis spectral shifts, and a shift in nu(CO) from 1648 to 1592 cm(-1) upon formation of mono(L-lysinehydroxamato)tetra(aquo)iron(III) (Fe(H(2)L)(H(2)O)(4)(4+)). The stepwise formation of tris(L-lysinehydroxamato)iron(III) from Fe(H(2)O)(6)(3+) and H(3)L(2+) was characterized by spectrophotometric titration, and the values for log beta(1), log beta(2), and log beta(3) are 6.80(9), 12.4(2), and 16.1(2), respectively, at 25 degrees C and I = 2.0 M (NaClO(4)). Stopped-flow spectrophotometry was used to study the proton-driven stepwise ligand dissociation kinetics of tris(L-lysinehydroxamato)iron(III) at 25 degrees C and I = 2.0 M (HClO(4)/NaClO(4)). Defining k(n) and k(-n) as the stepwise ligand dissociation and association rate constants and n as the number of bound LyHA ligands, k(3), k(-3), k(2), k(-2), k(1), and k(-1) are 3.0 x 10(4), 2.4 x 10(1), 3.9 x 10(2), 1.9 x 10(1), 1.4 x 10(-1), and 1.2 x 10(-1) M(-1) s(-1), respectively. These rate and equilibrium constants are compared with corresponding constants for Fe(III) complexes of acetohydroxamic acid (AHA) and N-methylacetohydroxamic acid (NMAHA) in the form of a linear free energy relationship. The role of electrostatics in these complexation reactions to form the highly charged Fe(LyHA)(3)(6+) species is discussed, and an interchange mechanism mediated by charge repulsion is presented. The reduction potential for tris(L-lysinehydroxamato)iron(III) is -214 mV (vs. NHE), and a comparison to other hydroxamic acid complexes of Fe(III) is made through a correlation between E(1/2) and pFe.     

A kinetic study of the reactions of Fe+ with N2O, N2, O2, CO2 and H2O, and the ligand-switching reactions Fe+.X + Y --> Fe+.Y + X (X = N2, O2, CO2; Y = O2, H2O)

A series of reactions involving Fe(+) ions were studied by the pulsed laser ablation of an iron target, with detection of ions by quadrupole mass spectrometry at the downstream end of a fast flow tube. The reactions of Fe(+) with N(2)O, N(2) and O(2) were studied in order to benchmark this new technique. Extending measurements of the rate coefficient for Fe(+) + N(2)O from 773 K to 185 K shows that the reaction exhibits marked non-Arrhenius behaviour, which appears to be explained by excitation of the N(2)O bending vibrational modes. The recombination of Fe(+) with CO(2) and H(2)O in He was then studied over a range of pressure and temperature. The data were fitted by RRKM theory combined with ab initio quantum calculations on Fe(+).CO(2) and Fe(+).H(2)O, yielding the following results (120-400 K and 0-10(3) Torr). For Fe(+) + CO(2): k(rec,0) = 1.0 x 10(-29) (T/300 K)(-2.31) cm(6) molecule(-2) s(-1); k(rec,infinity) = 8.1 x 10(-10) cm(3) molecule(-1) s(-1). For Fe(+) + H(2)O: k(rec,0) = 5.3 x 10(-29) (T/300 K)(-2.02) cm(6) molecule(-2) s(-1); k(rec,infinity) = 2.1 x 10(-9) (T/300 K)(-0.41) cm(3) molecule(-1) s(-1). The uncertainty in these rate coefficients is determined using a Monte Carlo procedure. A series of exothermic ligand-switching reactions were also studied at 294 K: k(Fe(+).N(2) + O(2)) = (3.17 +/- 0.41) x 10(-10), k(Fe(+).CO(2) + O(2)) = (2.16 +/- 0.35) x 10(-10), k(Fe(+).N(2) + H(2)O) = (1.25 +/- 0.14) x 10(-9) and k(Fe(+).O(2) + H(2)O) = (8.79 +/- 1.30) x 10(-10) cm(3) molecule(-1) s(-1), which are all between 36 and 52% of their theoretical upper limits calculated from long-range capture theory. Finally, the role of these reactions in the chemistry of meteor-ablated iron in the upper atmosphere is discussed. The removal rates of Fe(+) by N(2), O(2), CO(2) and H(2)O at 90 km altitude are approximately 0.1, 0.07, 3 x 10(-4) and 1 x 10(-6) s(-1), respectively. The initially formed Fe(+).N(2) and Fe(+).O(2) are converted into the H(2)O complex at approximately 0.05 s(-1). Fe(+).H(2)O should therefore be the most abundant single-ligand Fe(+) complex in the mesosphere below 90 km.     

Process of destruction of large unilamellar vesicles by a nonionic detergent,octylglucoside, and size growth factor in vesicle formation from phospholipid–detergent mixed micelles

The process of vesicle solubilization and size growth by detergents, especially by octylglucoside, was examined in detail in order to elucidate the phenomena observed in the vesicle-to-micelle transition and to clarify the size-determining factor of vesicles prepared by removing detergent from phospholipid–detergent mixed micelles. In the vesicle solubilization process, when the detergent concentration in the vesicle membrane reached a critical value, the collapse of large unilamellar vesicles (LUV) into small unilamellar vesicles (SUV) was observed. This newly appeared SUV were named SUV*. The SUV* could be produced by adding an appropriate amount of detergent to the SUV prepared by an ultrasonication method so as to increase the concentration to a little over the critical value, such as, in the case of adding octylglucoside, a molar ratio of 1.0–1.1 to phospholipid in the membrane phase. The SUV* containing octylglucoside were fusible and grow time-dependently, but those containing sodium cholate were not fusible. On the basis of the SUV* data, the following problems were solved: the variety of the size of the vesicles prepared by detergent removal from mixed micelles composed of a phospholipid and different detergents, or by different removal methods; the complex appearance of turbidity or vesicle size observed in vesicle destruction and formation; the conflict between LUV and SUV in the partition behavior of detergent and the size change with addition of detergent.     

CHLOROPHYLL PHOTOSENSITIZED ELECTRON TRANSFER IN PHOSPHOLIPID VESICLE SYSTEMS: COMPARISON OF INSIDE-OUTSIDE ASYMMETRY BETWEEN LARGE AND SMALL UNILAMELLAR VESICLES*

Abstract— We have determined the chlorophyll triplet quenching efficiencies, the chlorophyll cation radical yields and the conversion efficiencies of chlorophyll triplet to radical in large and small unilamellar phosphatidylcholine vesicles (LUV and SUV, respectively) in the presence of electrically-charged electron acceptors (ferricyanide and oxidized cytochrome c) located in either the inner or outer aqueous compartments of the vesicles. Both types of vesicles displayed inside-outside asymmetry, although the properties were reversed. Triplet quenching in SUV was more efficient when ferricyanide was located within the vesicle interior, whereas the reverse was true in LUV. When ferricyanide was located on the outside of the vesicles, the extent of triplet quenching in LUV was about two times that in SUV and the amount of cation radical formed in LUV was about two times that in SUV. Under these conditions, the conversion efficiencies of chlorophyll triplet to radical were 12.2% for LUV and 8.5% for SUV. With cytochrome c as an electron acceptor in negatively charged vesicles (25 mol per cent dixhexadecylphosphate incorporated) similar results were obtained. Again, the triplet quenching and radical yield inside-outside asymmetry properties were reversed between the two types of vesicles, and radical formation efficiencies when cyt c was located outside the vesicles were higher in LUV (11.7%) than in SUV (4.2%). We conclude that the inside-outside asymmetric photochemical behavior of unilamellar phosphatidylcholine vesicles is influenced by factors in addition to the difference in radius of curvature between the inside and outside surfaces. It is suggested that transmembrane electrostatic potentials may be involved. Furthermore, in the present system the properties of LUV were more favorable to photochemical electron transfer product formation than those of SUV.     

A supramolecular receptor of diatomic molecules (O2, CO, NO) in aqueous solution

A per-O-methylated beta-cyclodextrin dimer, Py2CD, was conveniently prepared via two steps: the Williamson reaction of 3,5-bis(bromomethyl)pyridine and beta-cyclodextrin (beta-CD) yielding 2A,2'A-O-[3,5-pyridinediylbis(methylene)bis-beta-cyclodextrin (bisCD) followed by the O-methylation of all the hydroxy groups of the bisCD. Py2CD formed a very stable 1:1 complex (Fe(III)PCD) with [5,10,15,20-tetrakis(p-sulfonatophenyl)porphinato]iron(III) (Fe(III)TPPS) in aqueous solution. Fe(III)PCD was reduced with Na2S2O4 to afford the Fe (II)TPPS/Py2CD complex (Fe(II)PCD). Dioxygen was bound to Fe(II)PCD, the P(1/2)(O2) values being 42.4 +/- 1.6 and 176 +/- 3 Torr at 3 and 25 degrees C, respectively. The k(on)(O2) and k(off)(O2) values for the dioxygen binding were determined to be 1.3 x 10(7) M(-1) s(-1) and 3.8 x 10(3) s(-1), respectively, at 25 degrees C. Although the dioxygen adduct was not very stable (K(O2) = k(on)(O2)/k(off)(O2) = 3.4 x 10(3) M(-1)), no autoxidation of the dioxygen adduct of Fe(II)PCD to Fe(III)PCD was observed. These results suggest that the encapsulation of Fe (II)TPPS by Py2CD strictly inhibits not only the extrusion of dioxygen from the cyclodextrin cage but also the penetration of a water molecule into the cage. The carbon monoxide affinity of Fe(II)PCD was much higher than the dioxygen affinity; the P(1/2)(CO), k(on)(CO), k(off)(CO), and K(CO) values being (1.6 +/- 0.2) x 10(-2) Torr, 2.4 x 10(6) M(-1) s(-1), 4.8 x 10(-2) s(-1), and 5.0 x 10(7) M(-1), respectively, at 25 degrees C. Fe(II)PCD also bound nitric oxide. The rate of the dissociation of NO from (NO)Fe(II)PCD ((5.58 +/- 0.42) x 10(-5) s(-1)) was in good agreement with the maximum rate ((5.12 +/- 0.18) x 10(-5) s(-1)) of the oxidation of (NO)Fe(II)PCD to Fe(III)PCD and NO3(-), suggesting that the autoxidation of (NO)Fe(II)PCD proceeds through the ligand exchange between NO and O2 followed by the rapid reaction of (O2)Fe(II)PCD with released NO, affording Fe(II)PCD and the NO3(-) anion inside the cyclodextrin cage.     

Nitrogen-14 NMR Study on Solvent Exchange of the Octahedral Cobalt(II) Ion in Neat 1,3-Propanediamine and n-Propylamine at Various Temperatures and Pressures. Tetrahedral-Octahedral Equilibrium of the Solvated Cobalt(II) Ion in n-Propylamine As Studied by EXAFS and Electronic Absorption Spectroscopy

Solvated cobalt(II) ions in neat 1,3-propanediamine (tn) and n-propylamine (pa) have been characterized by electronic absorption spectroscopy and extended X-ray absorption fine structure (EXAFS) spectroscopy. The equilibrium between tetrahedral and octahedral geometry for cobalt(II) ion has been observed in a neat pa solution, but not in neat diamine solutions such as tn and ethylenediamine (en). The thermodynamic parameters and equilibrium constant at 298 K for the geometrical equilibrium in pa were determined to be DeltaH degrees = -36.1 +/- 2.3 kJ mol(-1), DeltaS degrees = -163 +/- 8 J mol(-1) K(-1), and K(298) = 6.0 x 10(-3) M(-2), where K = [Co(pa)(6)(2+)]/{[Co(pa)(4)(2+)][pa](2)}. The equilibrium is caused by the large entropy gain in formation of the tetrahedral cobalt(II) species. The solvent exchange of cobalt(II) ion with octahedral geometry in tn and pa solutions has been studied by the (14)N NMR line-broadening method. The activation parameters and rate constants at 298 K for the solvent exchange reactions are as follows: DeltaH() = 49.3 +/- 0.9 kJ mol(-1), DeltaS() = 25 +/- 3 J mol(-1) K(-1), DeltaV() = 6.6 +/- 0.3 cm(3) mol(-1) at 302.1 K, and k(298) = 2.9 x 10(5) s(-1) for the tn exchange, and DeltaH() = 36.2 +/- 1.2 kJ mol(-1), DeltaS() = 35 +/- 6 J mol(-1) K(-1), and k(298) = 2.0 x 10(8) s(-1) for the pa exchange. By comparison of the activation parameters with those for the en exchange of cobalt(II) ion, it has been confirmed that the kinetic chelate strain effect is attributed to the large activation enthalpy for the bidentate chelate opening and that the enthalpic effect is smaller in the case of the six-membered tn chelate compared with the five-membered en chelate.     

Kinetics and mechanism of iron(III) dissociation from the dihydroxamate siderophores alcaligin and rhodotorulic acid

The kinetics and mechanism of siderophore ligand dissociation from their fully chelated Fe(III) complexes is described for the highly preorganized cyclic tetradentate alcaligin and random linear tetradentate rhodotorulic acid in aqueous solution at 25 degrees C (Fe2L3 + 6H+ reversible 2 Fe3+ aq + 3 H2L). At siderophore:Fe(III) ratios where Fe(III) is hexacoordinated, kinetic data for the H(+)-driven ligand dissociation from the Fe2L3 species is consistent with a singly ligand bridged structure for both the alcaligin and rhodotorulic acid complexes. Proton-driven ligand dissociation is found to proceed via parallel reaction paths for rhodotorulic acid, in contrast with the single path previously observed for the linear trihydroxamate siderophore ferrioxamine B. Parallel paths are also available for ligand dissociation from Fe2(alcaligin)3, although the efficiency of one path is greatly diminished and dissociation of the bis coordinated complex Fe(alcaligin)(OH2)2+ is extremely slow (k = 10(-5) M-1 s-1) due to the high degree of preorganization in the alcaligin siderophore. Mechanistic interpretations were further confirmed by investigating the kinetics of ligand dissociation from the ternary complexes Fe(alcaligin)(L) in aqueous acid where L = N-methylacetohydroxamic acid and glycine hydroxamic acid. The existence of multiple ligand dissociation paths is discussed in the context of siderophore mediated microbial iron transport.     

Atmospheric chemistry of hydrofluoroethers: Reaction of a series of hydrofluoroethers with OH radicals and Cl atoms, atmospheric lifetimes, and global warming potentials

The kinetics of the OH radical and Cl atom reactions with nine fluorinated ethers have been studied by the relative rate method at 298 K and 1013 hPa using gas chromatography-mass spectroscopy (GC-MS) detection: k(OH + CH3CH2OCF3) = (1.55 +/- 0.25) x 10(-13), k(OH + CF3CH2OCH3) = (5.7 +/- 0.8) x 10(-13),k(OH + CF3CH2OCHF2) = (9.1 +/- 1.1) x 10(-15), k(OH + CF3CHFOCHF2) = (6.5 +/- 0.8) x 10(-15), k(OH + CHF2CHFOCF3) = (6.8 +/- 1.1) x 10(-15), k(OH + CF3CHFOCF3) < 1 x 10(-15), k(OH + CF3CHFCF2OCHF2) = (1.69 +/- 0.26) x 10(-14), k(OH + CF3CHFCF2OCH2CH3) = (1.47 +/- 0.13) x 10(-13), k(OH + CF3CF2CF2OCHFCF3) < 1 x 10(-15), k(Cl + CH3CH2OCF3) = (2.2 +/- 0.8) x 10(-12), k(Cl + CF3CH2OCH3) = (1.8 +/- 0.9) x 10(-11), k(Cl + CF3CH2OCHF2) = (1.5 +/- 0.4) x 10(-14), k(Cl + CF3CHFOCHF2) = (1.1 +/- 1.9) x 10(-15), k(Cl + CHF2CHFOCF3) = (1.2 +/- 2.0) x 10(-15), k(Cl + CF3CHFOCF3) < 3 x 10(-15), k(Cl + CF3CHFCF2OCHF2) < 6 x 10(-16), k(Cl + CF3CHFCF2OCH2CH3) = (3.1 +/- 1.1) x 10(-12), and k(Cl + CF3CF2CF2OCHFCF3) < 3 x 10(-15) cm3 molecule(-1) s(-1). The error limits include three standard deviations (3 sigma) from the statistical data analyses, as well as the errors in the rate coefficients of the reference compounds that are used. Infrared absorption cross sections and estimates of the trophospheric lifetimes and the global warming potentials of the fluorinated ethers are presented. The atmospheric degradation of the compounds is discussed.     

Monitoring lipid membrane translocation of sodium dodecyl sulfate by isothermal titration calorimetry

We establish high-sensitivity isothermal titration calorimetry (ITC) as a fast, reliable, and versatile tool for assessing membrane translocation of charged compounds. A combination of ITC uptake and release titrations can discriminate between the two extreme cases of half-sided binding and complete transbilayer equilibration on the experimental time scale. To this end, we derive a general fit function for both assays that allows for incorporation of different membrane partitioning models. Electrostatic effects are taken into account with the aid of Gouy-Chapman theory, thus rendering uptake and release experiments amenable to the investigation of charged solutes. This is exemplified for the flip-flop of the anionic detergent sodium dodecyl sulfate (SDS) across the membranes of 100-nm-diameter unilamellar vesicles composed of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) in aqueous solution (10 mM phosphate buffer, 154 mM NaCl, pH 7.4). If repulsive electrostatic forces are accounted for adequately, SDS binding to POPC membranes can be evaluated on the basis of ideal mixing in all phases. At 25 degrees C, the intrinsic partition coefficient between the interfacial aqueous phase and the membrane amounts to 3.5 x 10(6); however, detergent flip-flop is negligibly slow under these conditions. Raising the temperature to 65 degrees C lowers the intrinsic partition coefficient to 1.4 x 10(6) but enables rapid transbilayer distribution of the detergent and, therefore, binding to or desorption from both membrane leaflets. Thus, combining a surface partition equilibrium with simple electrostatic theory appears highly useful in monitoring transmembrane movement of ionic compounds by ITC, thereby eliminating the need for specific reporter groups.     

When does an intermediate become a transition state? Degenerate isomerization without competing racemization during solvolysis of (S)-1-(3-nitrophenyl)ethyl tosylate

(S)-1-(3-Nitrophenyl)ethyl tosylate [(S)-2-OTs] was prepared in >99% enantiomeric excess and the change in the chiral purity of this compound was monitored during solvolysis in 50:50 trifluoroethanol/water. The barely detectable formation of 0.5% (R)-2-OTs after two half times for the solvolysis reaction was used to calculate a rate constant of k(rac) approximately equal to 4 x 10-6 s-1. This is 80-fold smaller than kiso = 3.2 x 10-4 s-1 for the isomerization that exchanges oxygen-16 and oxygen-18 of 3-NO2C6H413CH(Me)OS(18O)2Tos during solvolysis and 10-fold smaller than the minimum value of k(rac) = 4.6 x 10-5 s-1 predicted if isomerization and racemization products form by partitioning of a common ion-pair intermediate of a stepwise reaction. It is concluded that the isomerization reaction proceeds mainly by a pathway that avoids formation of this putative intermediate. It is suggested that the solvolysis reaction of 2-OTs may proceed by a stepwise preassociation mechanism where solvent "reorganization" precedes substrate ionization to form an ion-pair intermediate.     

NMR study of ligand exchange and electron self-exchange between oxo-centered trinuclear clusters [Fe3(μ3-O)(μ-O2CR)6(4-R'py)3](+/0)

The syntheses, single crystal X-ray structures, and magnetic properties of the homometallic μ?-oxo trinuclear clusters [Fe?(μ?-O)(μ-O?CCH?)?(4-Phpy)?](ClO?) (1) and [Fe?(μ?-O)(μ-O?CAd)?(4-Mepy)?](NO?) (2) are reported (Ad = adamantane). The persistence of the trinuclear structure within 1 and 2 in CD?Cl? and C?D?Cl? solutions in the temperature range 190-390 K is demonstrated by 1H NMR. An equilibrium between the mixed pyridine clusters [Fe?(μ?-O)(μ-O?CAd)?(4-Mepy)(3-x)(4-Phpy)(x)](NO?) (x = 0, 1, 2, 3) with a close to statistical distribution of these species is observed in CD?Cl? solutions. Variable-temperature NMR line-broadening made it possible to quantify the coordinated/free 4-Rpy exchanges at the iron centers of 1 and 2: k(ex)2?? = 6.5 ± 1.3 × 10?1 s?1, ΔH(?) = 89.47 ± 2 kJ mol?1, and ΔS(?) = +51.8 ± 6 J K?1 mol?1 for 1 and k(ex)2?? = 3.4 ± 0.5 × 10?1 s?1, ΔH(?) = 91.13 ± 2 kJ mol?1, and ΔS(?) = +51.9 ± 5 J K?1 mol?1 for 2. A limiting D mechanism is assigned for these ligand exchange reactions on the basis of first-order rate laws and positive and large entropies of activation. The exchange rates are 4 orders of magnitude slower than those observed for the ligand exchange on the reduced heterovalent cluster [Fe(III)?Fe(II)(μ?-O)(μ-O?CCH?)?(4-Phpy)?] (3). In 3, the intramolecular Fe(III)/Fe(II) electron exchange is too fast to be observed. At low temperatures, the 1/3 intermolecular second-order electron self-exchange reaction is faster than the 4-Phpy ligand exchange reactions on these two clusters, suggesting an outer-sphere mechanism: k?2?? = 72.4 ± 1.0 × 103 M?1 s?1, ΔH(?) = 18.18 ± 0.3 kJ mol?1, and ΔS(?) = -90.88 ± 1.0 J K?1 mol?1. The [Fe?(μ?-O)(μ-O?CCH?)?(4-Phpy)?](+/0) electron self-exchange reaction is compared with the more than 3 orders of magnitude faster [Ru?(μ?-O)(μ-O?CCH?)?(py)?](+/0) self-exchange reaction (ΔΔG(exptl)(?298) = 18.2 kJ mol?1). The theoretical estimated self-exchange rate constants for both processes compare reasonably well with the experimental values. The equilibrium constant for the formation of the precursor to the electron-transfer and the free energy of activation contribution for the solvent reorganization to reach the electron transfer step are taken to be the same for both redox couples. The larger ΔG(exptl)(?298) for the 1/3 iron self-exchange is attributed to the larger (11.1 kJ mol?1) inner-sphere reorganization energy of the 1 and 3 iron clusters in addition to a supplementary energy (6.1 kJ mol?1) which arises as a result of the fact that each encounter is not electron-transfer spin-allowed for the iron redox couple.     

Sulfate as a synergistic anion facilitating iron binding by the bacterial transferrin FbpA: the origins and effects of anion promiscuity

The ferric binding protein, FbpA, has been demonstrated to facilitate the transport of naked Fe3+ across the periplasmic space of several Gram-negative bacteria. The sequestration of iron by FbpA is facilitated by the presence of a synergistic anion, such as phosphate or sulfate. Here we report the sequestration of Fe3+ by FbpA in the presence of sulfate, at an assumed periplasmic pH of 6.5 to form FeFbpA-SO4 with K'(eff) = 1.7 x 10(16) M(-1) (at 20 degrees C, 50 mM MES, 200 mM KCl). The iron affinity of the FeFbpA-SO4 protein assembly is 2 orders of magnitude lower than when bound with phosphate and is the lowest of any of the FeFbpA-X assemblies yet reported. Iron reduction at the cytosolic membrane receptor may be an essential aspect of the periplasmic iron-transport process, and with an E(1/2) of -158 mV (NHE), FeFbpA-SO4 is the most easily reduced of all FeFbpA-X assemblies yet studied. The variation of FeFbpA-X assembly stability (K'(eff)) and ease of reduction (E(1/2)) with differing synergistic anions X(n-) are correlated over a range of 14 kJ, suggesting that the variations in redox potentials are due to stabilization of Fe3+ in FeFbpA-X by X(n-). Anion promiscuity of FbpA in the diverse composition of the periplasmic space is illustrated by the ex vivo exchange kinetics of FeFbpA-SO4 with phosphate and arsenate, where first-order kinetics with respect to FeFbpA-SO4 (k = 30 s(-1)) are observed at pH 6.5, independent of entering anion concentration and identity. Anion lability and influence on the iron affinity and reduction potential for FeFbpA-X support the hypothesis that synergistic anion exchange may be an important regulator in iron delivery to the cytosol. This structural and thermodynamic analysis of anion binding in FeFbpA-X provides additional insight into anion promiscuity and importance.     

A kinetic study of the reactions FeO+ + O, Fe+.N2 + O, Fe+.O2 + O and FeO+ + CO: implications for sporadic E layers in the upper atmosphere

These gas-phase reactions were studied by pulsed laser ablation of an iron target to produce Fe(+) in a fast flow tube, with detection of the ions by quadrupole mass spectrometry. Fe(+).N(2) and Fe(+).O(2) were produced by injecting N(2) and O(2), respectively, into the flow tube. FeO(+) was produced from Fe(+) by addition of N(2)O, or by ligand-switching from Fe(+).N(2) following the addition of atomic O. The following rate coefficients were measured: k(FeO(+) + O --> Fe(+) + O(2), 186-294 K) = (3.2 +/- 1.5) x 10(-11); k(Fe(+).N(2) + O --> FeO(+)+ N(2), 294 K) = (4.6 +/- 2.5) x 10(-10); k(Fe(+).O(2) + O --> FeO(+) + O(2), 294 K) = (6.3 +/- 2.7) x 10(-11); and k(FeO(+) + CO --> Fe(+) + CO(2), 294 K) = (1.59 +/- 0.34) x 10(-10) cm(3) molecule(-1) s(-1), where the quoted uncertainties are a combination of the 1sigma standard errors in the kinetic data and the systematic experimental errors. The surprisingly slow reaction between FeO(+) and O is examined using ab initio quantum calculations of the relevant potential energy surfaces. The importance of this reaction for controlling the lifetime of sporadic E layers is then demonstrated using a model of the upper mesosphere and lower thermosphere.