Journal of the American Chemical Society, Dec 30, 2004
The synthetic peptide encompassing residues 106-126 (PrP106-126, KTNMKHMAGAAAAGAVVGGLG) of the hu... more The synthetic peptide encompassing residues 106-126 (PrP106-126, KTNMKHMAGAAAAGAVVGGLG) of the human prion protein was considered for its binding properties toward copper(II), manganese(II) and zinc(II) at pH 5.7. 1H and 13C 1D spectra, 1H spin-lattice relaxation rates, and 1H-15N and 1H-13C HSQC 2D experiments were obtained in the absence and in the presence of metal ions. While Zn(II) was found to yield negligible effects upon any NMR parameter, metal-peptide association was demonstrated by the paramagnetic effects of Cu(II) and Mn(II) upon 1D and 2D spectra. Delineation of structures of metal complexes was sought by interpreting the paramagnetic effect on 1H spin-lattice relaxation rates. Exchange of peptide molecules from the metal coordination sphere was shown to provide sizable contribution to the observed relaxation rates. Such contribution was calculated in the case of Cu(II); whereas the faster paramagnetic rates of peptide molecules bound to Mn(II) were determining spin-lattice relaxation rates almost exclusively dominated by exchange. Proton-metal distances were therefore evaluated in the case of the Cu(II) complex only and used as restraints in molecular dynamics calculations where from the structure of the complex was obtained. The peptide was shown to bind copper through the imidazole nitrogen and the ionized amide nitrogen of His-111 and the amino-terminal group with the terminal carboxyl stabilizing the coordination sphere through ionic interactions. The data were interpreted as to demonstrate that the hydrophobic C-terminal region was not affecting the copper-binding properties of the peptide and that this hydrophobic tail is left free to interact with other target molecules. As for the complex with Mn(II), qualitative information was obtained on carbonyl oxygens of Gly-124 and Leu-125, beyond the terminal Gly-126 carboxyl, being at close distance from the metal ion, that also interacts, most likely, through a hydrogen bond of metal-bound water, with the imidazole ring of His-111.
Abstract A panel of ionic liquids has been synthesized and their effect on the vanadyl acetylacet... more Abstract A panel of ionic liquids has been synthesized and their effect on the vanadyl acetylacetonate solubility in acetonitrile has been firstly assessed. 1-Butyl-3-methylimidazolium acetate showed an unprecedented result, increasing the VO(acac)2 solubility in acetonitrile of more than one magnitude order (from 0.06 M to 1.1 M) opening new interesting horizons for the possible applications of this vanadium complex. The electrochemical effect of the considered ionic liquids has been subsequently investigated through cyclic voltammetry and linear sweep voltammetry with rotating disk electrode, determining diffusion coefficient and kinetic current of VO(acac)2 in the considered media. In order to achieve a deeper understanding on the examined systems, VO(acac)2 solutions in acetonitrile ILs were eventually studied through IR, UV–vis, and EPR spectroscopies, finding evidences, corroborated by DFT studies, of the formation of strong adducts between VO(acac)2 and ILs.
The thermodynamic potential of many reaction intermediates of redox enzymes allows the transfer o... more The thermodynamic potential of many reaction intermediates of redox enzymes allows the transfer of their electrons to oxygen as an alternative substrate. Membranes act as a physical barrier against the diffusion of oxygen and its reactive species derivatives (ROS); the intensity of the effect is, however, highly specific. For O2, the barrier effect is so low that it can be considered nonexistent. If at all, O2 movement is controlled at the lipid–water interface and this control is enhanced at high temperature and, in more rigid membranes, due to accumulation of cholesterol (Moller et al. 2005). For practical purposes, biological membranes do not represent any barrier for oxygen. Transfer of O•−2 is energetically unfavorable, since it is a charged species and needs to lose its hydration shell to cross membranes. However, cells have specific channels to transport biologically required ions (O•−2 included) across membranes (Lynch and Fridovich 1978). H2O2 is a small noncharged hydrophilic molecule which is transported across membranes through specific aquaporins (Bienert et al. 2007). HO• is the most reactive of the partially reduced oxygen species and reacts with phospholipids in the lipid bilayer, first with the polar heads which are inert to other ROS and, afterwards,with the acyl chains. Thus, membranes pose a physical barrier for the diffusion of HO• through reaction with the lipid or protein components of the bilayer. Organisms may alter the composition of their membranes in a controlled fashion to reduce permeability as observed in Saccharomyces cerevisiae (Folmer et al. 2008)
The multicopper oxidase from the hyperthermophilic archaeon Pyrobaculum aerophilum (McoP) was ove... more The multicopper oxidase from the hyperthermophilic archaeon Pyrobaculum aerophilum (McoP) was overproduced in Escherichia coli and purified to homogeneity. The enzyme consists of a single 49.6 kDa subunit, and the combined results of UV–visible, CD, EPR and resonance Raman spectroscopies showed the characteristic features of the multicopper oxidases. Analysis of the McoP sequence allowed its structure to be derived by comparative modeling methods. This model provided a criterion for designing meaningful site‐directed mutants of the enzyme. McoP is a hyperthermoactive and thermostable enzyme with an optimum reaction temperature of 85 °C, a half‐life of inactivation of ∼ 6 h at 80 °C, and temperature values at the midpoint from 97 to 112 °C. McoP is an efficient metallo‐oxidase that catalyzes the oxidation of cuprous and ferrous ions with turnover rate constants of 356 and 128 min−1, respectively, at 40 °C. It is noteworthy that McoP follows a ping‐pong mechanism, with three‐fold higher catalytic efficiency when using nitrous oxide as electron acceptor than when using dioxygen, the typical oxidizing substrate of multicopper oxidases. This finding led us to propose that McoP represents a novel archaeal nitrous oxide reductase that is most probably involved in the final step of the denitrification pathway of P. aerophilum.
Journal of Molecular Catalysis B-enzymatic, May 1, 2010
Laccase, isolated from Cerrena unicolor, is able to transform 3-amino-4-hydroxybenzensulfonic aci... more Laccase, isolated from Cerrena unicolor, is able to transform 3-amino-4-hydroxybenzensulfonic acid into a water soluble phenoxazine dye with an extinction coefficient (epsilon) of 8600 M-1 cm(-1). The dye has been characterized using a variety of different analytic and spectroscopic techniques like UV-vis spectroscopy, HPLC (High Performance Liquid Chromatography), ESI/MS (Electrospray Ionization Mass Spectrometry) and the following NMR experiments: H-1, C-13, TOCSY (Total Correlation Spectroscopy), HSQC (Heteronuclear Single Quantum Coherence). HMBC (Heteronuclear Multiple Bond Coherence) showing the structure of 2-amino-3-oxo-3H-phenoxazine-8-sulfonic acid. The advantages of the presented biocatalytic system, in alignment with chemical system to obtain Curie_22, are eco-sustainability and one step performance. (C) 2010 Elsevier B.V. All rights reserved.
New photocatalytic fibers made of sulfonated polyetheretherketone (SPEEK)/polypropylene (PP) are ... more New photocatalytic fibers made of sulfonated polyetheretherketone (SPEEK)/polypropylene (PP) are melt compounded and melt spun, first on laboratory scale and then on a semi-industrial scale. Fiber ...
Glycated human serum albumin (gHSA) undergoes conformational changes and unfolding events caused ... more Glycated human serum albumin (gHSA) undergoes conformational changes and unfolding events caused by free radicals. The glycation process results in a reduced ability of albumin to act as an endogenous scavenger and transporter protein in diabetes mellitus type 2 (T2DM) patients. Astaxanthin (ASX) in native form and complexed with metal ions (Cu2+ and Zn2+) has been shown to prevent gHSA from experiencing unfolding events. Furthermore, it improves protein stability of gHSA and human serum albumin (HSA) as it is shown through molecular dynamics studies. In this study, the ASX/ASX-metal ion complexes were reacted with both HSA/gHSA and analyzed with electronic paramagnetic resonance (EPR) spectroscopy, rheology and zeta sizer (particle size and zeta potential) analysis, circular dichroism (CD) spectroscopy and UV-Vis spectrophotometer measurements, as well as molecular electrostatic potential (MEP) and molecular docking calculations. The addition of metal ions to ASX improves its abili...
A library of five hybrids and six dimers of dihydroartemisinin and artesunic acid has been synthe... more A library of five hybrids and six dimers of dihydroartemisinin and artesunic acid has been synthetized in a stereo‐controlled manner and evaluated for the anticancer activity against metastatic melanoma cell line (RPMI7951). Among novel derivatives, three artesunic acid dimers showed antimelanoma activity and cancer selectivity, being not toxic on normal human fibroblast (C3PV) cell line. Among the three dimers, the one bearing 4‐hydroxybenzyl alcohol as a spacer showed no cytotoxic effect (CC50>300 μM) and high antimelanoma activity (IC50=0.05 μM), which was two orders of magnitude higher than that of parent artesunic acid, and of the same order of commercial drug paclitaxel. In addition, this dimer showed cancer‐type selectivity towards melanoma compared to prostate (PC3) and breast (MDA‐MB‐231) tumors. The occurrence of a radical mechanism was hypothesized by DFO and EPR analyses. Qualitative structure activity relationships highlighted the role of artesunic acid scaffold in the control of toxicity and antimelanoma activity.
Journal of the American Chemical Society, Dec 30, 2004
The synthetic peptide encompassing residues 106-126 (PrP106-126, KTNMKHMAGAAAAGAVVGGLG) of the hu... more The synthetic peptide encompassing residues 106-126 (PrP106-126, KTNMKHMAGAAAAGAVVGGLG) of the human prion protein was considered for its binding properties toward copper(II), manganese(II) and zinc(II) at pH 5.7. 1H and 13C 1D spectra, 1H spin-lattice relaxation rates, and 1H-15N and 1H-13C HSQC 2D experiments were obtained in the absence and in the presence of metal ions. While Zn(II) was found to yield negligible effects upon any NMR parameter, metal-peptide association was demonstrated by the paramagnetic effects of Cu(II) and Mn(II) upon 1D and 2D spectra. Delineation of structures of metal complexes was sought by interpreting the paramagnetic effect on 1H spin-lattice relaxation rates. Exchange of peptide molecules from the metal coordination sphere was shown to provide sizable contribution to the observed relaxation rates. Such contribution was calculated in the case of Cu(II); whereas the faster paramagnetic rates of peptide molecules bound to Mn(II) were determining spin-lattice relaxation rates almost exclusively dominated by exchange. Proton-metal distances were therefore evaluated in the case of the Cu(II) complex only and used as restraints in molecular dynamics calculations where from the structure of the complex was obtained. The peptide was shown to bind copper through the imidazole nitrogen and the ionized amide nitrogen of His-111 and the amino-terminal group with the terminal carboxyl stabilizing the coordination sphere through ionic interactions. The data were interpreted as to demonstrate that the hydrophobic C-terminal region was not affecting the copper-binding properties of the peptide and that this hydrophobic tail is left free to interact with other target molecules. As for the complex with Mn(II), qualitative information was obtained on carbonyl oxygens of Gly-124 and Leu-125, beyond the terminal Gly-126 carboxyl, being at close distance from the metal ion, that also interacts, most likely, through a hydrogen bond of metal-bound water, with the imidazole ring of His-111.
Abstract A panel of ionic liquids has been synthesized and their effect on the vanadyl acetylacet... more Abstract A panel of ionic liquids has been synthesized and their effect on the vanadyl acetylacetonate solubility in acetonitrile has been firstly assessed. 1-Butyl-3-methylimidazolium acetate showed an unprecedented result, increasing the VO(acac)2 solubility in acetonitrile of more than one magnitude order (from 0.06 M to 1.1 M) opening new interesting horizons for the possible applications of this vanadium complex. The electrochemical effect of the considered ionic liquids has been subsequently investigated through cyclic voltammetry and linear sweep voltammetry with rotating disk electrode, determining diffusion coefficient and kinetic current of VO(acac)2 in the considered media. In order to achieve a deeper understanding on the examined systems, VO(acac)2 solutions in acetonitrile ILs were eventually studied through IR, UV–vis, and EPR spectroscopies, finding evidences, corroborated by DFT studies, of the formation of strong adducts between VO(acac)2 and ILs.
The thermodynamic potential of many reaction intermediates of redox enzymes allows the transfer o... more The thermodynamic potential of many reaction intermediates of redox enzymes allows the transfer of their electrons to oxygen as an alternative substrate. Membranes act as a physical barrier against the diffusion of oxygen and its reactive species derivatives (ROS); the intensity of the effect is, however, highly specific. For O2, the barrier effect is so low that it can be considered nonexistent. If at all, O2 movement is controlled at the lipid–water interface and this control is enhanced at high temperature and, in more rigid membranes, due to accumulation of cholesterol (Moller et al. 2005). For practical purposes, biological membranes do not represent any barrier for oxygen. Transfer of O•−2 is energetically unfavorable, since it is a charged species and needs to lose its hydration shell to cross membranes. However, cells have specific channels to transport biologically required ions (O•−2 included) across membranes (Lynch and Fridovich 1978). H2O2 is a small noncharged hydrophilic molecule which is transported across membranes through specific aquaporins (Bienert et al. 2007). HO• is the most reactive of the partially reduced oxygen species and reacts with phospholipids in the lipid bilayer, first with the polar heads which are inert to other ROS and, afterwards,with the acyl chains. Thus, membranes pose a physical barrier for the diffusion of HO• through reaction with the lipid or protein components of the bilayer. Organisms may alter the composition of their membranes in a controlled fashion to reduce permeability as observed in Saccharomyces cerevisiae (Folmer et al. 2008)
The multicopper oxidase from the hyperthermophilic archaeon Pyrobaculum aerophilum (McoP) was ove... more The multicopper oxidase from the hyperthermophilic archaeon Pyrobaculum aerophilum (McoP) was overproduced in Escherichia coli and purified to homogeneity. The enzyme consists of a single 49.6 kDa subunit, and the combined results of UV–visible, CD, EPR and resonance Raman spectroscopies showed the characteristic features of the multicopper oxidases. Analysis of the McoP sequence allowed its structure to be derived by comparative modeling methods. This model provided a criterion for designing meaningful site‐directed mutants of the enzyme. McoP is a hyperthermoactive and thermostable enzyme with an optimum reaction temperature of 85 °C, a half‐life of inactivation of ∼ 6 h at 80 °C, and temperature values at the midpoint from 97 to 112 °C. McoP is an efficient metallo‐oxidase that catalyzes the oxidation of cuprous and ferrous ions with turnover rate constants of 356 and 128 min−1, respectively, at 40 °C. It is noteworthy that McoP follows a ping‐pong mechanism, with three‐fold higher catalytic efficiency when using nitrous oxide as electron acceptor than when using dioxygen, the typical oxidizing substrate of multicopper oxidases. This finding led us to propose that McoP represents a novel archaeal nitrous oxide reductase that is most probably involved in the final step of the denitrification pathway of P. aerophilum.
Journal of Molecular Catalysis B-enzymatic, May 1, 2010
Laccase, isolated from Cerrena unicolor, is able to transform 3-amino-4-hydroxybenzensulfonic aci... more Laccase, isolated from Cerrena unicolor, is able to transform 3-amino-4-hydroxybenzensulfonic acid into a water soluble phenoxazine dye with an extinction coefficient (epsilon) of 8600 M-1 cm(-1). The dye has been characterized using a variety of different analytic and spectroscopic techniques like UV-vis spectroscopy, HPLC (High Performance Liquid Chromatography), ESI/MS (Electrospray Ionization Mass Spectrometry) and the following NMR experiments: H-1, C-13, TOCSY (Total Correlation Spectroscopy), HSQC (Heteronuclear Single Quantum Coherence). HMBC (Heteronuclear Multiple Bond Coherence) showing the structure of 2-amino-3-oxo-3H-phenoxazine-8-sulfonic acid. The advantages of the presented biocatalytic system, in alignment with chemical system to obtain Curie_22, are eco-sustainability and one step performance. (C) 2010 Elsevier B.V. All rights reserved.
New photocatalytic fibers made of sulfonated polyetheretherketone (SPEEK)/polypropylene (PP) are ... more New photocatalytic fibers made of sulfonated polyetheretherketone (SPEEK)/polypropylene (PP) are melt compounded and melt spun, first on laboratory scale and then on a semi-industrial scale. Fiber ...
Glycated human serum albumin (gHSA) undergoes conformational changes and unfolding events caused ... more Glycated human serum albumin (gHSA) undergoes conformational changes and unfolding events caused by free radicals. The glycation process results in a reduced ability of albumin to act as an endogenous scavenger and transporter protein in diabetes mellitus type 2 (T2DM) patients. Astaxanthin (ASX) in native form and complexed with metal ions (Cu2+ and Zn2+) has been shown to prevent gHSA from experiencing unfolding events. Furthermore, it improves protein stability of gHSA and human serum albumin (HSA) as it is shown through molecular dynamics studies. In this study, the ASX/ASX-metal ion complexes were reacted with both HSA/gHSA and analyzed with electronic paramagnetic resonance (EPR) spectroscopy, rheology and zeta sizer (particle size and zeta potential) analysis, circular dichroism (CD) spectroscopy and UV-Vis spectrophotometer measurements, as well as molecular electrostatic potential (MEP) and molecular docking calculations. The addition of metal ions to ASX improves its abili...
A library of five hybrids and six dimers of dihydroartemisinin and artesunic acid has been synthe... more A library of five hybrids and six dimers of dihydroartemisinin and artesunic acid has been synthetized in a stereo‐controlled manner and evaluated for the anticancer activity against metastatic melanoma cell line (RPMI7951). Among novel derivatives, three artesunic acid dimers showed antimelanoma activity and cancer selectivity, being not toxic on normal human fibroblast (C3PV) cell line. Among the three dimers, the one bearing 4‐hydroxybenzyl alcohol as a spacer showed no cytotoxic effect (CC50>300 μM) and high antimelanoma activity (IC50=0.05 μM), which was two orders of magnitude higher than that of parent artesunic acid, and of the same order of commercial drug paclitaxel. In addition, this dimer showed cancer‐type selectivity towards melanoma compared to prostate (PC3) and breast (MDA‐MB‐231) tumors. The occurrence of a radical mechanism was hypothesized by DFO and EPR analyses. Qualitative structure activity relationships highlighted the role of artesunic acid scaffold in the control of toxicity and antimelanoma activity.
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