Interactions of Ru(II) polypyridyl complexes with DNA mismatches and abasic sites.

Polypyridyl based ruthenium(II) complexes, [Ru(bpy)2(furphen)](PF6)2 (1) and [Ru(bpy)2(imiphen)](PF6)2 (2) {furphen: 2-(furan-2-yl)-1H-imidazo[4,5-f][1,10]phenanthroline and imiphen: 2-(1H-imidazol-2-yl)-1H-imidazo[4,5-f][1,10]phenanthroline} were synthesized and characterized by ESI-MS, NMR, UV-Visible and fluorescence spectroscopic techniques. The interaction of Ru(II) complexes with calf-thymus DNA (CT DNA) as well as oligonucleotides containing mismatches and abasic sites was studied along with unmodified control DNA. Based on absorption titration studies, binding constants (Kb) for the interaction of complexes 1 and 2 with DNA were found to be 6.7 ± 0.2 × 10(3) and 4.9 ± 0.2 × 10(4) M(-1), respectively. Hydrodynamic studies revealed weak interactions between the two complexes and CT-DNA. Luminescence studies revealed that both the complexes exhibit a five-fold increase in emission upon addition of CT-DNA. The integrated emission intensity of complexes 1 and 2 with CC mismatch oligonucleotides was 1.5 and 1.2 fold higher than that of control GC match oligonucleotides, respectively. Both the complexes did not show any specificity towards abasic or other mismatch sites except for CC mismatch. The results from this study provide an insight into the requirements of ligand shape in recognising DNA mutations such as mismatch and selectivity between DNA mismatches.

A microscopic evaluation of collagen-bilirubin interactions: in vitro surface phenomenon.

This study is carried out to understand the morphology variations of collagen I matrices influenced by bilirubin. The characteristics of bilirubin interaction with collagen ascertained using various techniques like XRD, CLSM, fluorescence, SEM and AFM. These techniques are used to understand the distribution, expression and colocalization patterns of collagen-bilirubin complexes. The present investigation mimic the in vivo mechanisms created during the disorder condition like jaundice. Fluorescence technique elucidates the crucial role played by bilirubin deposition and interaction during collagen organization. Influence of bilirubin during collagen fibrillogenesis and banding patterns are clearly visualize using SEM. As a result, collagen-bilirubin complex provides different reconstructed patterns because of the influence of bilirubin concentration. Selectivity, specificity and spatial organization of collagen-bilirubin are determined through AFM imaging. Consequently, it is observed that the morphology and quantity of the bilirubin binding to collagen varied by the concentrations and the adsorption rate in protein solutions. Microscopic studies of collagen-bilirubin interaction confirms that bilirubin influence the fibrillogenesis and alter the rate of collagen organization depending on the bilirubin concentration. This knowledge helps to develop a novel drug to inhibit the interface point of interaction between collagen and bilirubin.

Microviscosity-induced conformational transition in ß-lactoglobulin in the presence of an ionic liquid.

This study reports on the helix-beta conformation transition of bovine ß-lactoglobulin (ßLG) prepared at two different pH conditions (pH 4 and 7.5) and in the presence of the ionic liquid 1-ethyl-3-methylimidazolium ethyl sulfate (IL-emes). The investigation was carried out by combining a range of techniques such as circular dichroic (CD) spectroscopy, steady-state fluorescence spectroscopy, isothermal titration calorimetry (ITC), and transmission electron microscopy. The influence of microviscosity induced by IL-emes on the secondary structure of ßLG was studied using a quartz crystal microbalance and correlated with the steady-state fluorescence emission. The effect of heat on the helix-beta transition in ßLG was directly measured by ITC by titrating ßLG with IL-emes. The net effect of heat after subtraction of the heat of dilution was negative in both cases, suggesting that the protein moves to a stable conformation. The changes in the overall aggregated structures were confirmed by transmission electron microscopy, where a shift in the size and morphology of aggregates was found, from large clusters (size of 70 nm) at pH 4 to smaller aggregates (size of 20 nm) at pH 7.5, which reduced to 7 nm in the presence of the IL. The transformation of helical to beta structure at pH 4 show that the folding pathway in the presence of the ionic liquid is hierarchical, whereas at neutral pH, it appeared to be nonhierarchical and the final native structure was acquired by nonlocal interactions through typical forces involved in the stabilization of the tertiary structure.

Reversible and irreversible conformational transitions in myoglobin: role of hydrated amino acid ionic liquid.

Hydrated phenylalanine ionic liquid (Phe-IL) has been used to solubilize myoglobin (Mb). Structural stability of Mb in Phe-IL analyzed using fluorescence and circular dichroism spectroscopy shows that for low levels of hydration of Phe-IL there is a large red shift in the fluorescence emission wavelength and the protein transforms to complete ß sheet from its native helical conformation. Rehydration or dilution reverses the ß sheet to an α helix which on aging organizes to micrometer-sized fibrils. At concentrations higher than 200 µM, the protein changes from ß to a more random coiled structure. Organization of the protein in Phe-IL in a Langmuir film at the air/water interface has been investigated using the surface pressure-molecular area isotherm and shows nearly the same surface tension for both pure Mb and Mb in Phe-IL. Scanning electron microscopy of the films of Mb in Phe-IL transferred using the Langmuir-Blodgett film technique show layered morphology. This study shows that the conformation of Mb is completely reversible going from ß â†’ helix → ß sheet up to 200 µM of Phe-IL. Similar surface tension values for Mb in water and in Phe-IL suggests that direct ion binding interactions with the protein coupled with the change in local viscosity from the IL seems to not only alter the secondary structure of individual proteins but also drives the self-assembly of the protein molecules leading finally to fibril formation.

Influence of sequence on the self-assembly of peptide nanoribbons on silicon substrates.

This work reports the formation of stable nanoassemblies of short pentapeptides LKLKL (pepI) and their mutated sequence LKKLL (pepII) obtained from their Langmuir-Blodgett films transferred onto hydrophilic and hydrophobic silicon substrates. The adsorption and assembly of the LB films of these peptides on solid surfaces have been studied by quartz crystal microbalance, surface plasmon resonance, and scanning electron microscopy. Both pepI and pepII assemble into nanosized ribbons, with diameters around 20-25 nm and lengths greater than 5 µm on hydrophobic surface, and tend to aggregate on hydrophilic surfaces with pepII showing twisted structures. Circular dichroic spectra of the films on a hydrophobic surface showed formation of a ß-sheet-like structure, while the corresponding solution spectra did not show any specific secondary structure. Our results demonstrate the formation of a two-dimensional dense array of nanoassemblies with either vertical or horizontal patterns from such short peptides that may find application in nanotechnology.

Preparation and antimicrobial activity of scleraldehyde from Schizophyllum commune.

The present study investigates the antimicrobial activity of oxidized schizophyllan (scleraldehyde) against Gram-positive and Gram-negative bacteria by diffusion and tube dilution analysis. Schizophyllan is a natural polysaccharide produced by fungi of the genus Schizophyllum. Periodate oxidation specifically cleaves the vicinal glycols in scleraldehyde to form their dialdehyde derivatives. The antibacterial activity exhibited by scleraldehyde was defined using various tests such as the disc diffusion assay, minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC). MIC and MBC values were found to be in the range of 3.0-8.0 mg/mL. Hence, the present studies establish that the scleraldehyde possesses effective antibacterial properties and can be used as a biopreservative for preservation of raw hides and skins.

Oxidation of benzaldehyde in films by Oxo-Cr(V) salen derivatives at solid/liquid interface.

The oxidation of benzaldehyde as a film on a solid surface by various substituted oxochromium(V) salen in solution has been studied by monitoring the change in contact angle of the oxidant at the film/liquid interface utilizing a Teflon cell of known hydrodynamics and controlled convection/diffusion. The kinetics of the redox reaction in bulk has been monitored by measuring the change in absorbance of the oxidant solution. The interfacial study permits analysis of adsorption of the oxidant followed by the oxidation of the substrate under pseudo-first-order conditions. A comparison of the independent surface-averaged kinetic data with those obtained in the solution phase oxidation reaction is made and a model is presented for the mechanism of the interfacial reaction. The kinetic investigation shows that the rate of oxidation is accelerated in the presence of an electron-withdrawing group and is faster at the solid/liquid interface compared to the bulk. The probable mechanism of the redox reaction is discussed.

Chromium(III)-mediated structural modification of glycoprotein: impact of the ligand and the oxidants.

The interaction of three types of chromium(III) complexes, [Cr(salen) (H2O2]+, [Cr(en)3]3+, and [Cr(EDTA) (H2O)]- with AGP has been investigated. [Cr(salen) (H2O2]+, [Cr(en)3]3+ and [Cr(EDTA) (H2O]- bind to Human alpha1-acid glycoprotein with a protein:metal ratio of 1:8, 1:6, and 1:4, respectively. The binding constant, K(b) was estimated to be 1.37 +/- 0.12 x 10(5) M(-1), 1.089 +/- 0.05 x 10(5) M(-1) and 5.3 +/- 0.05 x 10(4) M(-1) for [Cr(salen) (H2O2]+, [Cr(en)3]3+, and [Cr(EDTA) (H2O)]-, respectively. [Cr(en)3]3+ has been found to induce structural transition of AGP from the native twisted beta sheet to a more compact alpha-helix. The complexes, [Cr(salen) (H2O2]+ and [Cr(EDTA) (H2O]-, in the presence of H2O2, have been found to bring about nonspecific cleavage of AGP, whereas [Cr(en)3]3+ does not bring about any protein damage. Treatment of [Cr(salen) (H2O)2]+-protein adduct with iodosyl benzene on the other hand led to site specific cleavage of the protein. These results clearly demonstrate that protein damage brought about by chromium(III) complexes depends on the nature of the coordinated ligand, nature of the metal complex, and the nature of the oxidant.

Chromium removal and reduction in COD of tannery effluents by actinomycetes.

Tannery effluents are highly polluting and contain chroomium and high Cod and Bod. Alkalotolerant/alkalophilic actinomycetes NCIM 5080 and NCIM 5142 have been shown earlier to tolerate and accumulate chromium during growth also produce alkaline protease in presence of chromium ions, these properties of the isolates are suitable for treatment of tannery effluents which are alkaline and contain chromium and proteinaceous matter, both the actinomycetes are able to grow in undiluted tannery effluents and remove chromium almost completely and reduce the COD by 70%-80% during growth as well as by pregrown biomass.

Interaction of DNA with [Cr(Schiff base)(H(2)O)(2)]ClO(4).

The binding of Schiff base complexes of chromium(III) of the type [Cr(salen)(H(2)O)(2)](+) and [Cr(salprn)(H(2)O)(2)](+), where salen denotes 1,2-bis(salicylideneamino)ethane and salprn denotes 1,3-bis(salicylideneamino)propane to calf thymus DNA has been investigated by absorption, emission, circular dichroism, melting temperature and viscosity measurements. These chromium(III) complexes showed absorption hyperchromicity accompanied by red shift in charge transfer band, fluorescence enhancement, increase in melting temperature, some structural changes in CD spectra and changes in specific viscosity when bound to calf thymus DNA. The binding constant K(b) has been determined from absorption measurements for both the complexes and found to be (2.5+/-0. 4)x10(3) M(-1) for [Cr(salen)(H(2)O)(2)](+) and (1.7+/-0.3)x10(4) M(-1) for [Cr(salprn)(H(2)O)(2)](+). From the binding stoichiometry of DNA-[Cr(salprn)(H(2)O)(2)](+), the number of binding site size has been determined and found to be ten base pairs per bound complex molecule. The chromium(III) complexes also bring about single strand cleavage in plasmid DNA. The experimental results show that the chromium(III) complexes bind to DNA by non-intercalative mode. Major groove binding is the preferred mode of interaction for these Schiff base complexes of chromium(III).

DNA cleavage by a Chromium(III) complex.

A new Cr(III) complex with the empirical formula [Cr(Schiff base) (H(2)O)(2)]ClO(4), where the Schiff base is 2, 3-bis¿[(2-hydroxy-4-diethylamino) (phenyl) (methylene)]amino¿2-butenedinitrile has been synthesized and characterized spectroscopically. Binding of this complex to DNA has been studied using UV-visible spectroscopy. The complex has been found to bind to the major groove of DNA with a binding constant, K = (1.3 +/- 0.2) x 10(3) M(-1). The induced CD spectrum of the complex in the presence of DNA is also indicative of major groove binding. Gel electrophoresis of plasmid DNA in the presence of the complex shows that the complex brings about nicking of the DNA.

Protein degradation by peroxide catalyzed by chromium (III): role of coordinated ligand.

In order to understand the role of coordinated ligands in controlling the biotoxicity of chromium (III), interactions of three types of chromium (III) complexes viz. trans-diaquo [1,2 bis (salicyledeneamino) ethane chromium (III) perchlorate, [(Cr(salen)(H(2)O)(2)](ClO(4)); tris (ethylenediamine) chromium (III) chloride, [Cr(en)(3)]Cl(3), and monosodium ethylene diamine tetraacetato monoaquo chromiate (III), [Cr(EDTA)(H(2)O)]Na with BSA has been investigated. Spectroscopic and equilibrium dialysis studies show that the two cationic complexes Cr(salen)(H(2)O)(+)(2) and Cr(en)(3+)(3) bind to the protein with a protein-metal ratio of 1:8 and 1:4. The anionic complex Cr(EDTA)(H(2)O)(-) binds to the protein with a protein-metal ratio of 1:2. The binding constant K(b) as estimated from the fluorescence quenching studies has been found to be 7.6 +/- 0.4 x 10(3) M(-1), 3.1 +/- 0.2 x 10(2) M(-1), and 1.8 +/- 0.2 x 10(2) M(-1) for Cr(salen)(H(2)O)(+)(2), Cr(en)(3+)(3), and Cr(EDTA)(H(2)O)(-) respectively indicating that the thermodynamic stability of protein-chromium complex is Cr(salen)(H(2)O)(+)(2) > Cr(en)(3+)(3) approximately Cr(EDTA)(H(2)O)(-). The complexes Cr(salen)(H(2)O)(+)(2) and Cr(EDTA)(H(2)O)(-) in the presence of hydrogen peroxide have been found to bring about protein degradation, whereas Cr(en)(3+)(3) does not bring about any protein damage. This clearly shows that the nature of the chromium (III) complex plays a major role in the biotoxicity of chromium (III).

Chromium(III) hydrolytic oligomers: their relevance to protein binding.

The nature of chromium(III) complexes has been found to show a profound influence in its interaction with collagen. The hydrothermal stability of rat tail tendon (RTT) fibres treated with dimeric, trimeric and tetrameric species of chromium(III) has been found to be 102, 87 and 68 degrees C, while that of native RTT is 62 degrees C. This shows that the efficiency of crosslinking of collagen by chromium(III) species is dimeric > trimeric > tetrameric. This order of stabilisation is again confirmed by cyanogen bromide (CNBr) cleavage of RTT collagen treated with dimeric, trimeric and tetrameric chromium(III) species. CNBr has been found to cleave the collagen treated with tetrameric chromium(III) species extensively. On the other hand, dimer-treated collagen does not undergo any cleavage on CNBr treatment. The equilibrium constants for the reaction of a nucleophile like NCS(-) to the dimeric, trimeric and tetrameric species of chromium(III) have been found to be 15.7+/-0.1, 14.6+/-0.1 and 1.2+/-0.1 M(-1), respectively. These equilibrium constant values reflect the relative thermodynamic stability of the chromium(III) species-nucleophile complex. The low stabilising effect of the tetrameric species can be traced to its low thermodynamic affinity for nucleophiles.

Interaction of Schiff base with bovine serum albumin: site-specific photocleavage.

A Schiff-base ligand with donor/acceptor substituents viz. 2, 3-bis¿[(2-hydroxy-4-diethylamino) (phenyl) (methylene)]amino¿-2-butenedinitrile was synthesized, its binding properties with bovine serum albumin (BSA) and its site-specific photocleavage in the presence of cobaltous chloride have been evaluated. The Schiff-base ligand showed increase in absorption with a 5-nm red shift in the absorption maximum consistent with the binding of Schiff-base ligand to hydrophobic sites on the protein. The binding plot obtained from the absorption titration gives a binding constant of 6.4 +/- 0.3 x 10(4) M(-1). The CD spectrum of BSA in presence of the ligand shows that binding of the ligand leads to a change in the helicity of the protein. This ligand has been found to induce site-specific photocleavage of the protein in the presence of cobaltous chloride. The gel electrophoresis pattern of a photolyzed sample of BSA/Schiff-base ligand/cobaltous chloride shows that protein is cleaved into two polypeptide fragments, indicating site-specific binding for the ligand to the protein.

Chromium(III) interactions with nucleosides and nucleotides: a mass spectrometric study.

Interactions of [Cr(salprn)(H(2)O)(2)]ClO(4) with nucleosides and dinucleotides were studied using electrospray ionization mass spectrometry. The nucleosides 2'-deoxycytidine, thymidine, 2'-deoxyadenosine, 2'-deoxyguanosine, cytidine, adenosine and guanosine form 1 : 1 and 2 : 1 adducts with [Cr(salprn)](+), whereas the dinucleotides CpG, GpC, ApT, TpA and TpC form only the 1 : 1 adducts. Collisional activation (CA) spectra of these adducts reveal that Cr(+) attaches to the bases in nucleosides and to both the phosphate and base, especially cytosine and guanine moieties, in the nucleotides. The sugar residues appear to offer no binding sites as elimination of sugar residues is fairly abundant in the CA spectra of the adducts of many of the nucleosides.

Chromium(III) induced abnormalities in human lymphocyte cell proliferation: evidence for apoptosis.

The occupational hazards and respiratory problems of workers associated with the chromium related industries like mining, electroplating and tanning have received much focus. Although the cytotoxic and mutagenic effects of common chromium(VI) compounds are now established, to the best of our knowledge, this is the first report of apoptosis (programmed cell death) caused by a set of Cr(III) complexes in human lymphocyte cells. The effects of five Cr(III) complexes with differing ligand environment and structure as well as K2Cr2O7 on Phytohaemagglutinin (PHA) induced lymphocyte cell proliferation have been investigated. Two of the Cr(III) complexes and K2Cr2O7 are found to cause apoptosis of lymphocytes but not the others. A case for the importance of species specific effects rather than non specific metal oxidation state dependent processes has now been made.

Active site chemistry of lysyl oxidase.

The bimolecular reduction of the Cu(II)-based enzyme lysyl oxidase with two inorganic reductants, tris bipyridylchromium(II) and (1,3,6,8,10,13,16,19)-octaazabicyclo (6,6,6)eicosanecobalt(II) has been examined at various ionic strength and [H+] conditions. The electrochemical properties of the enzyme have also been examined. The results show that Cu(II) is the redox site in the enzyme and has E 1/2 = 0.05 +/- 0.005 V against SCE. The observed rate constants, kobs, for the reduction of the enzyme by either Cr(bpy)32+ or Co(sep)2+ at any concentration of the reductant increased with the ionic strength of the medium. The ionic strength dependence of kobs has been analyzed in terms of the charge of the active site being 1 +.