Hg(II)-coordination by sugar-acids: role of the hydroxy groups

A solution study on the ability of some derivatised sugars [glucuronic acid (GluA), galacturonic acid (GalA) and glucosaminic acid (GlNA)] to complex the Hg(II) ion is reported. The stability constants of the complex species were determined by potentiometric measurements while (1)H NMR experiments allow to define the coordination sites of sugar molecules. GluA coordinates the metal ion through the carboxylic oxygen and the O-4 hydroxyl group and is found to form more stable complexes with respect to GalA in which metal ligation is from the carboxylic oxygen and the O-5 ring oxygen. GlNA forms stable complexes chelating Hg(II) ion through carboxylic oxygen and the alpha-amino group. The ternary 2,2'-bipyridine containing systems were also investigated by means of potentiometric studies. The ML(2) complexes were also isolated in the solid state and characterised by IR spectroscopy.     

Cadmium tri-tert-butoxysilanethiolates: Structural and spectroscopic models of metal sites in proteins

Syntheses and crystal structures of two molecular, heteroleptic cadmium complexes with CdS₂NO₂ and CdS₂N₂ kernels are described. Bis(tri-tert-butoxysilanethiolate)(1-methylimidazole)cadmium(II) and bis(tri-tert-butoxysilanethiolate)bis(1-methylimidazole)cadmium(II) coexist at equilibrium in chloroform solutions with varying concentrations of bis[bis(tri-tert-butoxysilanethiolate)cadmium(II)] and 1-methylimidazole. The equilibrium is characterized by solution ¹¹³Cd NMR spectra. Solid state CP MAS ¹³C, ²⁹Si, ¹¹³Cd NMR data for the complexes are also reported, analyzed and compared with the results obtained for cadmium-substituted proteins. The similarities and differences between the structures of cadmium complexes and their zinc analogues are discussed.     

Preparation and photophysical properties of precursors of inorganic macromolecules. Mono and binuclear complexes of Ru(II) and terpyridine derivatized with thiophene and 4-(5-bromothiophene) groups

The preparation and characterization of mono and binuclear complexes of Ru(II) with a newly synthesized derivate of the terpyridine ligand, 4^′-(5-bromothiophene)-2,2^′,6^′,2″-terpyridine, are communicated. In the binuclear complex, 2,5-bis(2,2^′,6^′,2″-terpyridine-4^′yl)thiophene was used as a bridge between two Ru(II) centers. The new compounds were characterized by H NMR, UV-Vis and IR spectroscopies. Bands at ∼500 nm for the Ru(II) to terpyridine charge transfer transition and absorption bands at λ<400 nm assigned to intraligand transitions, π^*←π, centered in the tpy moiety were observed in the UV-Vis spectra of the complexes. Irradiation of the complexes in CH₃CN at 337 or 500 nm induced luminescence with maxima at ∼670 nm and lifetimes τ?10² ns. Time-resolved absorption spectroscopy revealed the formation of long-lived species during the decay of the metal to ligand charge transfer excited states. The intermediates were tentatively assigned as unstable products of ligand-substitution or orthometalation excited state reactions.     

Two-dimensional rotating-frame Overhauser spectroscopy (ROESY) and (13)C NMR study of the interactions between maltodextrin and an anionic surfactant

Rotational frame nuclear Overhauser effect spectroscopy (ROESY) and (13)C NMR measurements were carried out to study the molecular interaction between maltodextrin, a digestive byproduct of starch, and an anionic surfactant. Significant differences in chemical shifts were observed when sodium dodecyl sulfate (SDS) was introduced into the maltodextrin (DE 10) solutions. (13)C NMR measurement indicated that there were downfield shifts and broadening of peaks, especially in the region of 75-81 and 100-103 ppm, which were assigned to carbons 1 and 4 of the d-glucopyranose residues of maltodextrin, respectively. ROESY spectra indicated cross-peaks between the SDS and maltodextrin protons. These peaks can arise only in the case of the designated SDS protons and maltodextrin protons being less than 0.5 nm apart for a substantial period of time. The most intense cross-peaks are those between the central CH(2) protons of SDS near 1.2 ppm and the maltodextrin protons ranging from 3.5 to 3.9 ppm. The SDS-H3 CH(2) protons were resolved from the bulk of the SDS protons, with peaks and shoulders at 1.25 ppm, which indicated an especially strong interaction of the SDS hydrophobic tail with MD6 and some less intense interactions with MD2, 4, and 5.     

Complexes of sodium vanadate(V) with methyl alpha-D-mannopyranoside,methyl alpha- and beta-D-galactopyranoside,and selected O-methyl derivatives: a 51V and 13C NMR study

The hydroxyl group stereochemistry of complexation of sodium vanadate(V) with Me alpha-Manp, Me alpha- and beta-Galp and selected O-methyl derivatives in D(2)O was determined by 51V, 1D and 2D 13C NMR spectroscopy at pD 7.8. The 51V approach served to show the extent of complexation and the minimum number of esters formed. That of Me alpha-Manp gave rise mainly to a 51V signal at delta -515, identical with that of its 4,6-di-O-methyl derivative, which had only a 2,3-cis-diol exposed. The 13C NMR spectra contained much weaker signals of the complexes, but both glycosides showed strong C-2 and C-3 alpha-shifts of +17.3 and +10.8 ppm, respectively. As expected, Me 2,3-Me(2)-alpha-Manp, which contains a 4,6-diol, did not complex. Me Galp anomers and their derivatives showed more diversity in the structure of its oxyvanadium derivatives. Me alpha-Galp, with its 3,4-cis-diol, complexed to give rise to 51V signals at delta -495 (9%), -508 (10%), and -534 (4%). These shifts and proportions were maintained with Me beta-Galp and Me 6Me-alpha-Galp. 51V NMR spectroscopy showed that Me 3Me-beta-Galp, with its possibly available 4,6-diol, did not complex. Similarly, Me alpha-Galp+vanadate gave a 13C DEPT spectrum that did not contain an inverted signal at delta >71.4, as would be expected of a C-6 resonance suffering a strong downfield alpha-shift. Me 2,6-Me(2)-alpha-Galp, with a 3,4-cis-diol group, gave rise to two 51V signals of complexes at delta -492 (9%) and -508 (9%), showing more than one structure of oxyvanadium derivatives.     

Synthesis, characterization and DNA binding properties of oligopyridine-ruthenium(II)-amino acid conjugates

The DNA-binding properties of a number of ruthenium oligopyridine complexes with conjugated amino acids having the general formulae [Ru(terpy)(4-COY-4'-Mebpy)(X)](n)(+), X=NO (n=3), X=Cl (n=1) and NO(2) (n=1) and Y=AlaCONH(2) and TrpCONH(2) are reported. The new complexes were spectroscopically characterized and their DNA-binding properties were studied by means of circular dichroism (CD), (23)Na and (31)P NMR spectroscopy. The results show that the chlorido complexes interact by coordination to the DNA bases with the conjugated amino acid able to provide an additional interaction with the DNA helix. In addition, electrostatic interactions between all studied complexes and the DNA polyanion were observed. The nitro complexes were found to be insignificant, affecting only the (31)P NMR signal, probably due to changes in the hydration sphere of the DNA close to the phosphates.     

Regular oscillatory behavior of aqueous solutions of CuII salts related to effects on equilibrium dynamics of ortho/para hydrogen spin isomers of water

Cell surface and growth-related NADH oxidases with protein disulfide-thiol interchange activity, ECTO-NOX, exhibit copper-dependent, clock-related, temperature-independent and entrainable patterns of regular oscillations in the rate of oxidation of NAD(P)H as do aqueous solutions of copper salts. Because of time scale similarities, a basis for the oscillatory patterns in nuclear spin orientations of the hydrogen atoms of the copper-associated water was sought. Extended X-ray absorption fine structure (EXAFS) measurements at 9302 eV on pure water were periodic with a ca. 3.5 min peak to peak separation. Decomposition fits revealed 5 unequally spaced maxima similar to those observed previously for Cu(II)Cl(2) to generate a period length of about 18 min. With D(2)O, the period length was proportionately increased by 30% to 24 min. The redox potential of water and of D(2)O also oscillated with 18 and 24 min period lengths, respectively. Measurements in the middle infrared spectral region above a water sample surface revealed apparent oscillations in the two alternative orientations of the nuclear spins (ortho and para) of the hydrogen atoms of the water or D(2)O with 5 unequally spaced maxima and respective period lengths of 18 and 24 min. Thus, the time keeping oscillations of ECTO-NOX proteins appear to reflect the equilibrium dynamics of ortho-para hydrogen atom spin ratios of water where the presence of metal cations such as Cu(II) in solution determine period length.     

Synthesis and structure of cadmium and zinc complexes of dehydroacetic acid

The cadmium and zinc complexes of Dehydroacetic Acid (DHA) Zn(DHA)₂(H₂O)₂ and Cd(DHA)₂(H₂O)₂ were synthesized and the derivatives Zn(ADH)₂(DMSO)₂ and Cd(ADH)₂(DMSO)₂ were prepared through substitution of the water ligands by DMSO. To characterize structural differences between the Cd and Zn complexes, a series of analyses were carried out: ¹H, ¹³C, and ¹¹³Cd NMR in solution and ¹³C and ¹¹³Cd NMR in the solid state, infra-red spectra, thermo gravimetric analysis (TGA), differential calorimetric analysis (DSC) and elemental analysis (CHNO). The X-ray crystal structures of the complexes Zn(DHA)₂(DMSO)₂ and Cd(DHA)₂(DMSO)₂ are also reported. The coordination around the metal atoms in the solid state is best described as distorted octahedra. The two chelating DHA ligands define an equatorial plane and the axial positions are occupied by two monodentate DMSO ligands coordinated by oxygen atoms, in the trans,trans,trans configuration. Significant differences were found between the Cd and Zn coordination spheres, with the latter forming relatively looser octahedral complexes.     

Spectroscopic aspects of the coordination modes of 2,4-dithiohydantoins: Experimental and theoretical study on copper and nickel complexes of cyclohexanespiro-5-(2,4-dithiohydantoin)

The complexation of cyclohexanespiro-5-(2,4-dithiohydantoin), L, with copper and nickel was studied by means of experimental and theoretical methods. The Cu(I) and Ni(II) complexes were synthesized and characterized using ¹³C CPMAS NMR, IR and FAB-MS. Reduction of Cu(II) ions and the formation of Cu(I) complexes with dithiohydantoin was proved. Various coordination modes were investigated on the basis of calculated (DFT-GIAO) shielding constants of the free ligand and model structures of the complexes. General trends in the changes of spectroscopic parameters (NMR chemical shifts, vibrational modes) upon different types of coordination were outlined. Dimeric structures for the Cu(I) and Ni(II) complexes were proposed in which the ligands were coordinated in N3^S4- and N3^S2-bridging ways, respectively, acting as monoanions. The results demonstrate that the combined experimental (¹³C CPMAS NMR, IR) and theoretical (DFT) approach can be used to characterize the molecular structure of solid complexes for which crystallographic data are not available.     

Complexation of vanadium(V) oxyanions with hexopyranose- and mannopyranoseuronic acid-containing polysaccharides: stereochemical considerations

Carbohydrates containing galactopyranosyl and mannopyranosyl units with vicinal cis-diols were treated with NaVO(3) in D(2)O, and complexation was determined by (51)V NMR spectroscopy. Me alpha-Galp, Me beta-Galp (3,4-cis-diols), and Me alpha-Manp (2,3-cis-diol) complexed, but Me beta-Manp barely did so. This low degree of complexation also occurred with a beta-mannan containing alternate (1-->3)- and (1-->4)-linkages and an alginate having beta-ManpA blocks. In contrast, branched alpha-mannans complexed readily, although the (51)V resonances for one with side chains terminated with alpha-Manp-(1-->3)-alpha-Manp-(1--> differed from another with only alpha-Manp-(1-->2)-alpha-Manp-(1--> groups. The anomeric configuration of Me alpha-Galp and Me beta-Galp, each with 3,4-cis-diols remote from C-1, gave rise to three (51)V signals of complexes with similar shifts and proportions. The shifts of a galactomannan with terminal alpha-Galp-(1-->2)-alpha-Manp- were the same as those with alpha-Galp-(1-->6)-beta-Manp- groups, but fewer complexes were formed with the former structure, probably due to greater steric crowding of the vanadate esters. Most of the complexes gave rise to a signal in the delta515 region, consistent with the dimeric trigonal-bipyramidal structure.     

Formation of a new hybrid complex via coordination interaction between 5,10,15-tritolyl-20-(4- and 3-pyridyl)porphyrin or 5,10,15-triphenyl-20-(4-pyridyl)porphyrin and the α-[MSiW11O39] Keggin-type polyoxometalate (M = Co and Ni)

New complexes based on a coordination interaction between a pyridyl-porphyrin (namely 5,10,15-tritolyl-20-(4-pyridyl)porphyrin, 5,10,15-tritolyl-20-(3-pyridyl)porphyrin or 5,10,15-triphenyl-20-(4-pyridyl)porphyrin) and a Keggin-type polyoxometalate (α-[MSiW₁₁O₃₉]⁶⁻, M = Co²⁺ or Ni²⁺) are formed in solution. The formation of these complexes is clearly evidenced by steady-state and time-resolved luminescence measurements. A strong quenching of the porphyrin fluorescence, accompanied by an important shortening of the fluorescence lifetime, is observed upon addition of the POM and formation of the complexes. Using a variant of the Job’s method from the luminescence spectra, the association constants of the complexes have been estimated to be around 10⁶ L mol⁻¹. Paramagnetic ¹H NMR experiments confirm the formation of the complexes. Indeed, in addition to broadenings of the signals, the coordination binding of the POM to the porphyrin induces large high-frequency shifts for the protons of the pyridyl group coordinated to the paramagnetic metal, and low-frequency shifts for all the other resonances.     

Assignment Strategy for Fast Relaxing Signals: Complete Aminoacid Identification in Thulium Substituted Calbindin D9K

Paramagnetic proteins generally contain regions with diverse relaxation properties. Nuclei in regions far from the metal center may behave like those in diamagnetic proteins, but those closer to the metal experience rapid relaxation with accompanying line broadening. We have used a set of NMR experiments optimized to capture data from these various concentric regions in assigning the signals from a paramagnetic Calbindin D 9K derivative in which one of the two calcium ions has been replaced by thulium(III). Normal double- and triple-resonance experiments with 1H detection were used in collecting data from nuclei in the diamagnetic-like region; these approaches identified signals from fewer than 50% of the amino acid residues (those with d > 17.5 A from thulium(III)). Paramagnetism-optimized two-dimensional NMR experiments with 1H detection were used in collecting data from nuclei in the next nearer region (d > 15 A). Standard (d > 14 A) and optimized (d > 9 A) 13C direct-detection experiments were used to capture data from nuclei in the next layer. Finally nuclei closest to the metal were detected by one-dimensional 13C (d > 5 A) and one-dimensional 15N data collection (d > 4.2 A). NMR signals were assigned on the basis of through-bond correlations and, for signals closest to the metal, pseudocontact shifts. The latter were determined from chemical shift differences between assigned signals in thulium(III) and lanthanum(III) derivatives of Calbindin D 9K and they were interpreted on the basis of a structural model for the lanthanide-substituted protein. This approach yielded assignments of at least one resonance per amino acid residue, including those in the thulium(III) coordination sphere.     

1H, 113Cd, and 31P NMR of osteocalcin (bovine gamma-carboxyglutamic acid containing protein)

The 1H (500-MHz), 113Cd (44-MHz), and 31P (81-MHz) NMR spectra of the bovine gamma-carboxyglutamate- (Gla-) containing protein osteocalcin and its Ca(II) and Cd(II) complexes in solution have been obtained. The 1H NMR spectrum of the native protein shows narrow resonances and a highly resolved multiplet structure suggesting rotational freedom of the side chains. In comparison to the simulated 1H NMR spectrum of a random polypeptide chain of the same amino acid composition, there is moderate chemical shift dispersion, indicating some conformational restraints to be present. Ca(II) binding broadens all 1H resonances, so severely at four Ca(II) ions per molecule that few structural conclusions can be made. Cd(II) substituted for Ca(II) has the same effect, and 113Cd NMR shows the Cd(II) to be in intermediate chemical exchange on the chemical shift time scale. Estimates of the chemical exchange rates required for 1H and 113Cd line broadening suggest a range of Kd values for the metal ion complexes from 10(-6) M to as high as 10(-3) M depending on the number of metal ions bound. Alternatively, 1H line broadening could be explained by relatively slow conformational fluxes in the protein induced by labile metal ion binding to one or more sites. Cd(II) when used to form a cadmium-phosphate mineral analogous to hydroxylapatite results in a crystal lattice that removes osteocalcin from solution just as effectively as hydroxylapatite. 113Cd(II) exchange at the binding sites of osteocalcin in solution is slowed dramatically by the addition of HPO4(2-). 31P NMR shows the interaction of phosphate with the protein to require the metal ion.(ABSTRACT TRUNCATED AT 250 WORDS)     

Dynamic pictures of membrane proteins in two-dimensional crystal, lipid bilayer and detergent as revealed by site-directed solid-state 13C NMR

We have compared site-directed 13C solid-state NMR spectra of [3-13C]Ala- and/or [1-13C]Val-labeled membrane proteins, including bacteriorhodopsin (bR), pharaonis phoborhodopin (ppR), its cognate transducer (pHtrII) and Escherichia coli diacylglycerol kinase (DGK), in two-dimensional (2D) crystal, lipid bilayers, and detergent. Restricted fluctuation motions of these membrane proteins due to oligomerization of bR by specific protein-protein interactions in the 2D crystalline lattice or protein complex between ppR and pHtrII provide the most favorable environment to yield well-resolved, fully visible 13C NMR signals for [3-13C]Ala-labeled proteins. In contrast, several signals from such membrane proteins were broadened or lost owing to interference of inherent fluctuation frequencies (10(4)-10(5)Hz) with frequency of either proton decoupling or magic angle spinning, if their 13C NMR spectra were recorded as a monomer in lipid bilayers at ambient temperature. The presence of such protein dynamics is essential for the respective proteins to achieve their own biological functions. Finally, spectral broadening found for bR and DGK in detergents were discussed.     

Ni(II) complexes with Schiff bases derived from amino sugars

It was found by 1H and 13C NMR spectroscopy that the Schiff base, 2-deoxy-2-(2-hydroxybenzaldimino)-D-glucopyranose exhibits enol-imine-keto-amine and anomeric equilibria in methanolic, and in dimethyl sulfoxide solutions. The reaction of the Schiff base with nickel acetate gave the bidentate, mononuclear Ni(II) complex that was characterized by spectroscopic methods and by cyclic voltammetry. The coordination of the Schiff base to the metal is through the enol-imine tautomeric form, and the anomeric equilibrium remains in dimethyl sulfoxide solutions. This complex was also obtained by reaction of D-glucosamine with Ni(II) salicylaldehydate. The same reaction was employed for the synthesis of bis-N-[2-deoxy-D-galactopyranosyl-2-(2-hydroxybenzaldiminate)]Ni(II). The small paramagnetic shifts of the 1H NMR resonances of the complexes suggest that paramagnetic species are present in low proportions.     

Unique 31P Spectral Response to the Formation of a Specific Restriction Enzyme–DNA Complex

Protein-induced distortion is a dramatic but not universally observed feature of sequence-specific DNA interactions. This is illustrated by the crystal structures of restriction enzyme–DNA complexes: While some of these structures exhibit DNA distortion, others do not. Among the latter is PvuII endonuclease, a small enzyme that is also amenable to NMR spectroscopic studies. Here ³¹P NMR spectroscopy is applied to demonstrate the unique spectral response of DNA to sequence-specific protein interactions. The ³¹P NMR spectrum of a noncognate DNA exhibits only spectral broadening upon the addition of enzyme. However, when enzyme is added to target DNA, a number of ³¹P resonances shift dramatically. The magnitudes of the chemical shifts (2–3 ppm) are among the largest observed. Site-specific substitution with phosphoramidates and phosphorothioates are used analyze these effects. While such spectral features have been interpreted as indicative of DNA backbone distortions, FRET analysis indicates that this does not occur in PvuII-cognate DNA complexes in solution. The distinct ³¹P spectral signature observed for cognate DNA mirrors that observed for the enzyme, underscoring the unique features of cognate complex formation.     

Synthesis, spectroscopical characterization and the biological activity in vitro of new platinum(II) complexes with imidazo[1,5-a]-1,3,5-triazine derivatives and dimethylsulfoxide

A series of platinum(II) complexes with 6,8-dimethylimidazo[1,5-a]-1,3,5-triazin-4(3H)-one (6,8-DiMe-4-O-IMT) (I) and 6,8-dimethyl-2-thioxo-2,3-dihydroimidazo[1,5-a]-1,3,5-triazin-4(1H)-one (6,8-DiMe-4-O-2-S-IMT) (II) of formula trans-[PtCl₂(dmso)(6,8-DiMe-4-O-IMT)] (1a) and trans-[PtCl₂(dmso)(6,8-DiMe-4-O-2-S-IMT)] (2a) have been prepared and characterized with ¹H, ¹³C, ¹⁵N, ¹⁹⁵Pt NMR and IR. Significant ¹⁵N NMR upfield coordination shifts (81-96 ppm) of N(7) atom indicate this nitrogen atom as a coordination site. The multinuclear NMR and IR spectra indicate the square planar geometry with N(7) bonded heterocycles, S-bonded dimethylsulfoxide and two trans chloride anions. The platinum(II) complexes were tested for their antiproliferative activity in vitro against the cells of four human cell lines: SW707 rectal adenocarcinoma, A549 non-small cell lung carcinoma, T47D breast cancer and HCV29T bladder cancer. The activity of (1a, 2a) was lower than that of cisplatin.     

Complete 1H, 13C and 15N NMR assignments and secondary structure of the 269-residue serine protease PB92 from Bacillus alcalophilus

Summary The ¹H, ¹³C and ¹⁵N NMR resonances of serine protease PB92 have been assigned using 3D tripleresonance NMR techniques. With a molecular weight of 27 kDa (269 residues) this protein is one of the largest monomeric proteins assigned so far. The side-chain assignments were based mainly on 3D H(C)CH and 3D (H)CCH COSY and TOCSY experiments. The set of assignments encompasses all backbone carbonyl and CH_n carbons, all amide (NH and NH₂) nitrogens and 99.2% of the amide and CH_n protons. The secondary structure and general topology appear to be identical to those found in the crystal structure of serine protease PB92 [Van der Laan et al. (1992) Protein Eng., 5, 405–411], as judged by chemical shift deviations from random coil values, NH exchange data and analysis of NOEs between backbone NH groups.Abbreviations 2D/3D/4D two-/three-/four-dimensional - HSQC heteronuclear single-quantum coherence - HMQC heteronuclear multiple-quantum coherence - COSY correlation spectroscopy - TOCSY total correlation spectroscopy - NOE nuclear Overhauser enhancement (connectivity) - NOESY 2D NOE spectroscopy Experiment nomenclature (H(C)CH, etc.) follows the conventions used elsewhere [e.g. Ikura et al. (1990) Biochemistry, 29, 4659–4667].     

SOD-like activity of Mn(II) beta-octabromo-meso-tetrakis(N-methylpyridinium-3-yl)porphyrin equals that of the enzyme itself

Mn porphyrins are among the most efficient SOD mimics with potency approaching that of SOD enzymes. The most potent ones, Mn(III) N-alkylpyridylporphyrins bear positive charges in a close proximity to the metal site, affording thermodynamic and kinetic facilitation for the reaction with negatively charged superoxide. The addition of electron-withdrawing bromines onto beta-pyrrolic positions dramatically improves thermodynamic facilitation for the O2*- dismutation. We have previously characterized the para isomer, Mn(II)Br(8)TM-4-PyP(4+) [Mn(II) beta-octabromo-meso-tetrakis(N-methylpyridinium-4-yl)porphyrin]. Herein we fully characterized its meta analogue, Mn(II)Br(8)TM-3-PyP(4+) with respect to UV/vis spectroscopy, electron spray mass spectrometry, electrochemistry, O2*- dismutation, metal-ligand stability, and the ability to protect SOD-deficient Escherichia coli in comparison with its para analogue. The increased electron-deficiency of the metal center stabilizes Mn in its +2 oxidation state. The metal-centered Mn(III)/Mn(II) reduction potential, E((1/2))=+468 mV vs NHE, is increased by 416 mV with respect to non-brominated analogue, Mn(III)TM-3-PyP(5+) and is only 12 mV less positive than for para isomer. Yet, the complex is significantly more stable towards the loss of metal than its para analogue. As expected, based on the structure-activity relationships, an increase in E((1/2)) results in a higher catalytic rate constant for the O2*- dismutation, log k(cat)> or =8.85; 1.5-fold increase with respect to the para isomer. The IC(50) was calculated to be < or =3.7 nM. Manipulation of the electron-deficiency of a cationic porphyrin resulted, therefore, in the highest k(cat) ever reported for a metalloporphyrin, being essentially identical to the k(cat) of superoxide dismutases (log k(cat)=8.84-9.30). The positive kinetic salt effect points to the unexpected, unique and first time recorded behavior of Mn beta-octabrominated porphyrins when compared to other Mn porphyrins studied thus far. When species of opposing charges react, the increase in ionic strength invariably results in the decreased rate constant; with brominated porphyrins the opposite was found to be true. The effect is 3.5-fold greater with meta than with para isomer, which is discussed with respect to the closer proximity of the quaternary nitrogens of the meta isomer to the metal center than that of the para isomer. The potency of Mn(II)Br(8)TM-3-PyP(4+) was corroborated by in vivo studies, where 500 nM allows SOD-deficient E. coli to grow >60% of the growth of wild type; at concentrations > or =5 microM it exhibits toxicity. Our work shows that exceptionally high k(cat) for the O2*- disproportionation can be achieved not only with an N(5)-type coordination motif, as rationalized previously for aza crown ether (cyclic polyamines) complexes, but also with a N(4)-type motif as in the Mn porphyrin case; both motifs sharing "up-down-up-down" steric arrangement.     

Zinc(II) and cadmium(II) metal complexes of thiamine pyrophosphate and 2-(α-hydroxyethyl)thiamine pyrophosphate: models for activation of pyruvate decarboxylase

Metal complexes of thiamine pyrophosphate (TPP) of the general formula [M₂(TPPH)₂Cl₂].4H₂O (M =Zn²⁺, Cd²⁺) were isolated from methanolic solutions and characterized by elemental analysis, FT-IR, and multinuclear NMR spectroscopies. The data provide evidence for the bonding of the metals to the N(1') atom of the pyrimidine ring and to the pyrophosphate group. The stability constant measurements of TPP and 2-(α-hydroxyethyl)thiamine pyrophosphate (HETPP) metal complexes in aqueous solution imply the formation of dimeric complex species similar to the isolated solid products. They indicate also that HETPP forms more stable metal complexes than does TPP. To evaluate the coenzyme action of TPP and HETPP metal complexes, enzymic studies have been done using pyruvate decarboxylase apoenzyme. TPP metal complexes do not bind to the apoenzyme, unlike the Zn(II)-HETPP complex which can act as coenzyme. Considering these results, possible functional implications for thiamine involvement in catalysis are discussed. Received 13 September 1999 / Accepted 4 January 2000