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1.
Int J Mol Sci ; 22(22)2021 Nov 21.
Artigo em Inglês | MEDLINE | ID: mdl-34830424

RESUMO

Fusobacterium nucleatum is one of the most notorious species involved in colorectal cancer. It was reported that numerous outer membrane proteins (OMP) are actively involved in carcinogenesis. In this paper, the structure and stability of certain complexes, as well as DNA cleavage and ROS generation by fragments of OMP, were investigated using experimental and theoretical methods. Mass spectrometry, potentiometry, UV-Vis, CD, EPR, gel electrophoresis and calculations at the density functional theory (DFT) level were applied. Two consecutive model peptides, Ac-AKGHEHQLE-NH2 and Ac-FGEHEHGRD-NH2, were studied. Both of these were rendered to form a variety of thermodynamically stable complexes with copper(II) ions. All of the complexes were stabilized, mainly due to interactions of metal with nitrogen and oxygen donor atoms, as well as rich hydrogen bond networks. It was also concluded that these complexes in the presence of hydrogen peroxide or ascorbic acid can effectively produce hydroxyl radicals and have an ability to cleave the DNA strands. Surprisingly, the second studied ligand at the micromolar concentration range causes overall DNA degradation.

2.
J Inorg Biochem ; 224: 111586, 2021 11.
Artigo em Inglês | MEDLINE | ID: mdl-34425476

RESUMO

Fusobacterium nucleatum (F. nucleatum) is one of the most abundant Gram-negative anaerobic bacteria, part of the gut, and oral commensal flora, generally found in human dental plaque. Its presence could be associated with various human diseases, including, e.g., periodontal, angina, lung and gynecological abscesses. This bacteria can enter the blood circulation as a result of periodontal infection. It was proven that F. nucleatum migrates from its primary site of colonization in the oral cavity to other parts of the body. It could cause numerous diseases, including cancers. On the other hand, it was shown that Fusobacterium produces significant amounts of butyric acid, which is a great source of energy for colonocytes (anti-inflammatory cells). Therefore, it is very interesting to get to know the two faces of F. nucleatum.

3.
J Inorg Biochem ; 215: 111332, 2021 02.
Artigo em Inglês | MEDLINE | ID: mdl-33340803

RESUMO

It has been reported that numerous of Fusobacterium nucleatum outer membrane proteins take part in cancerogenesis. Therefore, it is very interesting to study their interactions with metal ions and the ability to produce reactive oxygen species, which may be involved in cancer progression. Since investigations of metal binding to proteins are often based on fragments that contain the metal-binding domains, designing model peptides should be very mindful. As was shown in this paper, very similar protein fragments may behave differentially. Herein, combined potentiometric, spectroscopic, and computational studies were performed to determine metal ion binding by ligands constituting fragments of porin protein P1. Two studied tetrapeptides (Ac-KEHK-NH2 and Ac-EHKA-NH2) that have common EHK motif have different coordination properties and reactivity. Therefore, we should be cautious when transferring the behavior of small peptide fragments to whole protein.


Assuntos
Proteínas da Membrana Bacteriana Externa/metabolismo , Complexos de Coordenação/química , Cobre/química , Fusobacterium nucleatum/metabolismo , Peptídeos/metabolismo , Porinas/metabolismo , Proteínas da Membrana Bacteriana Externa/química , Dicroísmo Circular/métodos , Clivagem do DNA , Peróxido de Hidrogênio/metabolismo , Ligantes , Neoplasias/metabolismo , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/metabolismo , Peptídeos/química , Porinas/química , Potenciometria/métodos , Espécies Reativas de Oxigênio/metabolismo
4.
J Inorg Biochem ; 213: 111275, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-33091731

RESUMO

Often, in the search for a highly defined scientific phenomenon, a different one becomes apparent. This was also the case of this work, in the scope of which we planned to search for metal-enhanced, novel antibacterial/antifungal compounds. Instead, we denied the existence of such and revealed the details of the bioinorganic chemistry of Zn(II)-alloferon complexes. Zinc(II) complexes of alloferon 1 and 2, ligands with a sequential difference of one amino acid only, show a substantially different coordination pattern at physiological pH. In the case of Zn(II)-alloferon 1 species, a histamine-like binding mode is observed (N-terminal amine and imidazole of His-1) and the coordination sphere is completed with the imidazole nitrogens of His-6 and His-9; His-12 is not involved in binding. In the case of Zn(II)-alloferon 2, the N-terminal amine and all the three imidazoles present in the sequence participate in the coordination, however, with the chemical shift of His-5 being less affected than those of other imidazoles. The histamine-like binding in Zn(II)-alloferon 1 complex strongly enhances its thermodynamic stability in comparison to the His-1 lacking alloferon 2 analogue. Despite previous reports on the antibacterial and antifungal activity of alloferon 1, no such activity was detected, neither for alloferon 1 and 2 nor for their Zn(II) complexes.


Assuntos
Complexos de Coordenação/química , Peptídeos/química , Zinco/química , Sequência de Aminoácidos , Antibacterianos/farmacologia , Complexos de Coordenação/farmacologia , Histidina/química , Ligantes , Espectrometria de Massas/métodos , Testes de Sensibilidade Microbiana , Peptídeos/farmacologia , Espectroscopia de Prótons por Ressonância Magnética/métodos , Relação Estrutura-Atividade , Termodinâmica
5.
Pharmaceuticals (Basel) ; 13(9)2020 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-32882888

RESUMO

Zn(II) is an inhibitor of SARS-CoV-2's RNA-dependent RNA polymerase, and chloroquine and hydroxychloroquine are Zn(II) ionophores-this statement gives a curious mind a lot to think about. We show results of the first clinical trials on chloroquine (CQ) and hydroxychloroquine (HCQ) in the treatment of COVID-19, as well as earlier reports on the anticoronaviral properties of these two compounds and of Zn(II) itself. Other FDA-approved Zn(II) ionophores are given a decent amount of attention and are thought of as possible COVID-19 therapeutics.

6.
J Nanobiotechnology ; 18(1): 95, 2020 Jul 13.
Artigo em Inglês | MEDLINE | ID: mdl-32660596

RESUMO

BACKGROUND: Currently, the diagnosis and treatment of neuroblastomas-the most frequent solid tumors in children-exploit the norepinephrine transporter (hNET) via radiolabeled norepinephrine analogs. We aim to develop a nanomedicine-based strategy towards precision therapy by targeting hNET cell-surface protein with hNET-derived homing peptides. RESULTS: The peptides (seq. GASNGINAYL and SLWERLAYGI) were shown to bind high-resolution homology models of hNET in silico. In particular, one unique binding site has marked the sequence and structural similarities of both peptides, while most of the contribution to the interaction was attributed to the electrostatic energy of Asn and Arg (< - 228 kJ/mol). The peptides were comprehensively characterized by computational and spectroscopic methods showing ~ 21% ß-sheets/aggregation for GASNGINAYL and ~ 27% α-helix for SLWERLAYGI. After decorating 12-nm ferritin-based nanovehicles with cysteinated peptides, both peptides exhibited high potential for use in actively targeted neuroblastoma nanotherapy with exceptional in vitro biocompatibility and stability, showing minor yet distinct influences of the peptides on the global expression profiles. Upon binding to hNET with fast binding kinetics, GASNGINAYLC peptides enabled rapid endocytosis of ferritins into neuroblastoma cells, leading to apoptosis due to increased selective cytotoxicity of transported payload ellipticine. Peptide-coated nanovehicles significantly showed higher levels of early apoptosis after 6 h than non-coated nanovehicles (11% and 7.3%, respectively). Furthermore, targeting with the GASNGINAYLC peptide led to significantly higher degree of late apoptosis compared to the SLWERLAYGIC peptide (9.3% and 4.4%, respectively). These findings were supported by increased formation of reactive oxygen species, down-regulation of survivin and Bcl-2 and up-regulated p53. CONCLUSION: This novel homing nanovehicle employing GASNGINAYLC peptide was shown to induce rapid endocytosis of ellipticine-loaded ferritins into neuroblastoma cells in selective fashion and with successful payload. Future homing peptide development via lead optimization and functional analysis can pave the way towards efficient peptide-based active delivery of nanomedicines to neuroblastoma cells.


Assuntos
Sistemas de Liberação de Medicamentos/métodos , Endocitose/genética , Nanoestruturas/química , Neuroblastoma/metabolismo , Proteínas da Membrana Plasmática de Transporte de Norepinefrina , Antineoplásicos/química , Antineoplásicos/farmacocinética , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Ferritinas/química , Humanos , Nanomedicina , Proteínas da Membrana Plasmática de Transporte de Norepinefrina/química , Proteínas da Membrana Plasmática de Transporte de Norepinefrina/genética , Proteínas da Membrana Plasmática de Transporte de Norepinefrina/metabolismo , Peptídeos/química , Peptídeos/genética , Peptídeos/metabolismo
7.
Future Microbiol ; 15: 259-271, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32271108

RESUMO

Aim: Characterization of the ability of Fusobacterium nucleatum DSM 15643 and DSM 20482 strains in the presence of Cu2+ and H2O2 to reactive oxygen species generation. Method: Spectrophotometric ABTS (2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) method was used. Results: Determination of: MIC for Cu2+, H2O2 and ABTS; survivability of F. nucleatum under atmospheric oxygen exposure; the level and rate constants of free radicals production by the bacteria. Conclusion: F. nucleatum in the presence of Cu2+ and H2O2 is able to generate free radicals. Reactive oxygen species are produced mainly outside the bacterial cell, which suggests that outer membrane proteins may be involved in oxidative process.


Assuntos
Fusobacterium nucleatum/química , Fusobacterium nucleatum/metabolismo , Espécies Reativas de Oxigênio/análise , Espécies Reativas de Oxigênio/metabolismo , Espectrofotometria/métodos , Benzotiazóis/química , Cobre/farmacologia , Fusobacterium nucleatum/efeitos dos fármacos , Peróxido de Hidrogênio/farmacologia , Ácidos Sulfônicos/química
8.
J Inorg Biochem ; 195: 71-82, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-30927561

RESUMO

Phleomycin is one of the anticancer glycopeptide antibiotics which cause DNA cleavage. It is commonly used as a copper(II) complex. Therefore, it is important to study the metal ion binding process and to define the coordination mode. In this paper, we describe the acid-base properties of phleomycin and the coordination sphere of the Cu(II) cation. In the metal binding process up to five nitrogen donor atoms can be involved. Four of them in the same plane deriving from: the pyrimidine ring, secondary amine of ß-aminoalanine, imidazole and amide of the nearest peptide bond (from ß-hydroxyhistidine) and in the apical position from the α-amino functional group of ß-aminoalanine, resulting complex has a square-pyramidal geometry. Phleomycin complexes are able to induce single- and double-stranded DNA damage when they are accompanied by one-electron reductants, such as dithiothreitol, glutathione, 2-mercaptoethanol or ascorbic acid. In such conditions they produce reactive oxygen species which are responsible for DNA cleavage. The metal ion binding site is relatively close to the nucleic acid interacting moiety. This supports the hypothesis that copper ion is important in the anticancer activity which involves DNA degradation.


Assuntos
Complexos de Coordenação/química , Clivagem do DNA , DNA/química , Fleomicinas/química , Ácido Ascórbico/química , Cobre/química , Quebras de DNA de Cadeia Simples , Teoria da Densidade Funcional , Ligação de Hidrogênio , Cinética , Modelos Químicos , Estrutura Molecular
9.
J Inorg Biochem ; 189: 69-80, 2018 12.
Artigo em Inglês | MEDLINE | ID: mdl-30243120

RESUMO

Fusobacterium nucleatum is an anaerobic, Gram-negative bacteria linked to colon cancer. It is interesting to determine how metal ions interact with bacterial adhesin proteins. To this end, the coordination of ATDAAS-NH2 and MKKFL-NH2 fragments of Fusobacterium adhesin A (FadA) to copper(II) ions was studied by potentiometry, spectroscopic techniques (UV-Vis, CD, EPR and NMR) and the density functional theory (DFT) methods. At pH 6.8 (colon physiological pH), the metal ion in the first peptide (ATDAAS-NH2) is coordinated by one oxygen and three nitrogen donors while in the second one (MKKFL-NH2) - by sulfur and three nitrogen atoms. Both complexes form two five- and one six-membered stable chelate rings. Moreover, reactivity studies confirmed the production of reactive oxygen species such as hydroxyl radical, superoxide anion radical and singlet oxygen. Generation of reactive oxygen species (ROS) was observed during gel electrophoresis and spectroscopic assays with reporting molecules like NDMA (N,N-dimethyl-p-nitrosoaniline) and NBT (Nitrotetrazolium Blue Chloride). All reactions were conducted in the presence of hydrogen peroxide as endogenous oxidant.


Assuntos
Adesinas Bacterianas/química , Cobre/química , Fusobacterium nucleatum/química , Espectroscopia de Ressonância de Spin Eletrônica , Concentração de Íons de Hidrogênio , Potenciometria , Espécies Reativas de Oxigênio/química , Superóxidos/química
10.
Bioconjug Chem ; 29(9): 2954-2969, 2018 09 19.
Artigo em Inglês | MEDLINE | ID: mdl-30086240

RESUMO

Novicidin (NVC), is a membrane-penetrating peptide, which forms a stable complex with Zn-Schiff base with interesting antitumor selectivity. We studied NVC derivatives to determine functional roles of key amino acids in toxicity, helicity, and binding of the Zn-Schiff base complex. Trimmed derivatives highlighted the role of peptide length and helicity in toxicity and membrane penetration. The removal of Lys from position 1 and 2 strongly increases the ability to disrupt the membranes. The trimming of the N-terminal residues significantly increases the stability of peptide helicity enhancing penetrating properties. Gly residue derivatives undermined a role of peptide bending in membrane penetration and toxicity. After the substitution of the central Gly derivatives with Ile or Lys, the peptides retained toxicity. These results illustrate the minor role of central helix bending in NVC toxicity. Binding-site-peptide derivatives identified His residue as the sole Zn-Schiff base binding site and eliminated the role of other aromatic residues.


Assuntos
Peptídeos Catiônicos Antimicrobianos/administração & dosagem , Sistemas de Liberação de Medicamentos , Bases de Schiff/química , Zinco/administração & dosagem , Sequência de Aminoácidos , Peptídeos Catiônicos Antimicrobianos/química , Peptídeos Catiônicos Antimicrobianos/toxicidade , Sítios de Ligação , Linhagem Celular Tumoral , Permeabilidade da Membrana Celular , Glicina/química , Humanos , Ligantes , Conformação Proteica , Espectroscopia de Prótons por Ressonância Magnética , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Espectroscopia de Infravermelho com Transformada de Fourier , Zinco/química
11.
Eur J Med Chem ; 143: 997-1009, 2018 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-29232589

RESUMO

Peptide antibiotics are produced by bacterial, mammalian, insect or plant organisms in defense against invasive microbial pathogens. Therefore, they are gaining importance as anti-infective agents. There are a number of antibiotics that require metal ions to function properly. Metal ions play a key role in their action and are involved in specific interactions with proteins, nucleic acids and other biomolecules. On the other hand, it is well known that some antimicrobial agents possess functional groups that enable them interacting with metal ions present in physiological fluids. Some findings support a hypothesis that they may alter the serum metal ions concentration in humans. Complexes usually have a higher positive charge than uncomplexed compounds. This means that they might interact more tightly with polyanionic DNA and RNA molecules. It has been shown that several metal ion complexes with antibiotics promote degradation of DNA. Some of them, such as bleomycin, form stable complexes with redox metal ions and split the nucleic acids chain via the free radicals mechanism. However, this is not a rule. For example blasticidin does not cause DNA damage. This indicates that some peptide antibiotics can be considered as ligands that effectively lower the oxidative activity of transition metal ions.


Assuntos
Antibacterianos/farmacologia , Compostos Organometálicos/farmacologia , Peptídeos/farmacologia , Elementos de Transição/farmacologia , Animais , Antibacterianos/síntese química , Antibacterianos/química , Humanos , Íons/química , Íons/farmacologia , Testes de Sensibilidade Microbiana , Estrutura Molecular , Compostos Organometálicos/síntese química , Compostos Organometálicos/química , Peptídeos/síntese química , Peptídeos/química , Elementos de Transição/química
12.
J Inorg Biochem ; 175: 167-178, 2017 10.
Artigo em Inglês | MEDLINE | ID: mdl-28755574

RESUMO

Goserelin acetate (Gos) as a synthetic analog of the mammalian gonadotropin-releasing hormone (GnRH) is widely used in the treatment of sex hormone-related conditions. In this paper we present the chemical and biological aspects of its interaction with Cu(II) ions. The mode of Cu(II) binding and the thermodynamic stability of the obtained complexes were characterized by potentiometry, UV-Vis and CD spectroscopic methods. The DFT calculations were applied in order to investigate and confirm the molecular structure of the studied systems. The experimental and theoretical results clearly indicated the involvement of three nitrogens from the peptide and two oxygens from the acetate moieties in the Cu(II) coordination under physiological conditions. The investigated metallopeptide caused single- and/or double cleavage of the sugar-phosphate backbone of the plasmid DNA in the reaction accompanied by endogenous substances such as hydrogen peroxide or ascorbic acid. The degradation of the DNA molecule occurred via the free radical mechanism. Calculations based on measured spectra allowed determining the kinetic parameters of OH formation. The cytotoxic effects of Gos and its metallo-derivative were tested in vitro towards two cancer cell lines (A549 - human lung adenocarcinoma, CT26 - mouse colon carcinoma).


Assuntos
Complexos de Coordenação , Citotoxinas , DNA de Neoplasias/química , Gosserrelina , Células A549 , Animais , Complexos de Coordenação/síntese química , Complexos de Coordenação/química , Complexos de Coordenação/farmacologia , Cobre , Citotoxinas/síntese química , Citotoxinas/química , Citotoxinas/farmacologia , DNA de Neoplasias/metabolismo , Gosserrelina/química , Gosserrelina/farmacologia , Humanos , Camundongos , Oxirredução
13.
ChemistryOpen ; 6(1): 46-56, 2017 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-28168150

RESUMO

Here, we describe the characterization of new RNA-cleaving DNAzymes that showed the highest catalytic efficiency at pH 4.0 to 4.5, and were completely inactive at pH values higher than 5.0. Importantly, these DNAzymes did not require any divalent metal ion cofactors for catalysis. This clearly suggests that protonated nucleic bases are involved in the folding of the DNAzymes into catalytically active structures and/or in the cleavage mechanism. The trans-acting DNAzyme variants were also catalytically active. Mutational analysis revealed a conservative character of the DNAzyme catalytic core that underpins the high structural requirements of the cleavage mechanism. A significant advantage of the described DNAzymes is that they are inactive at pH values close to physiological pH and under a wide range of conditions in the presence of monovalent and divalent metal ions. These pH-dependent DNAzymes could be used as molecular cassettes in biotechnology or nanotechnology, in molecular processes that consist of several steps. The results expand the repertoire of DNAzymes that are active under nonphysiological conditions and shed new light on the possible mechanisms of catalysis.

14.
J Sulphur Chem ; 37(3): 307-327, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-27516805

RESUMO

4-C-Alkyl/aryl-S-ribosylhomocysteine (SRH) analogues were prepared by coupling of homocysteine with 4-substituted ribofuranose derivatives. The diastereoselective incorporation of the methyl substituent into the 4 position of the ribose ring was accomplished by addition of methylmagnesium bromide to the protected ribitol-4-ulose yielding the 4-C-methylribitol in 85% yield as single 4R diastereomer. The 4-C hexyl, octyl, vinyl, and aryl ribitols were prepared analogously. Chelation controlled addition of a carbanion to ketones from the (Si-face) was responsible for the observed stereochemical outcome. Oxidation of the primary alcohol of the 4-C ribitols with the catalytic amount of tetrapropylammonium perruthenate in the presence of N-methylmorpholine N-oxide produced 4-C-alkylribono-1,4-lactones in high yields. Mesylation of the latter compounds at the 5-hydroxyl position and treatment with a protected homocysteine thiolate afforded protected 4-C-alkyl/aryl-SRH analogues as the lactones. Reduction with lithium triethylborohydride and successive global deprotections with TFA afforded 4-C-alkyl/aryl SRH analogues. These analogues might impede the S-ribosylhomocysteinase(LuxS)-catalyzed reaction by preventing ß-elimination of a homocysteine molecule, and thus depleting the production of quorum sensing signaling molecule AI-2.

15.
J Inorg Biochem ; 151: 67-74, 2015 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-26028475

RESUMO

Colistin and transition metal ions are commonly used as feed additives for livestock animals. This work presents the results of an analysis of combined potentiometric and spectroscopic (UV-vis, EPR, CD, NMR) data which lead to conclude that colistin is able to effectively chelate copper(II) ions. In cell-free system the oxidative activity of the complex manifests itself in the plasmid DNA destruction with simultaneous generation of reactive OH species, when accompanied by hydrogen peroxide or ascorbic acid. The degradation of RNA occurs most likely via a hydrolytic mechanism not only for complexed compound but also colistin alone. Therefore, huge amounts of the used antibiotic for nontherapeutic purposes might have a potential influence on livestock health.


Assuntos
Colistina/química , Cobre/química , Cobre/farmacologia , Ácidos Nucleicos/química , Sistema Livre de Células/efeitos dos fármacos , Dicroísmo Circular , Complexos de Coordenação , Íons/farmacologia , Estrutura Molecular , Oxirredução/efeitos dos fármacos
16.
Dalton Trans ; 44(17): 8138-49, 2015 May 07.
Artigo em Inglês | MEDLINE | ID: mdl-25836771

RESUMO

In vitro selection was performed to search for RNA-cleaving DNAzymes catalytically active with Cd(2+) ions from the oligonucleotide combinatorial library with a 23-nucleotide random region. All the selected, catalytically active variants turned out to belong to the 8-17 type DNAzyme. Three DNAzymes were prepared in shortened, cis-acting versions which were subjected to a detailed study of the kinetic properties and metal ion preferences. Although the selection protocol was designed for Cd(2+)-dependent DNAzymes, the variants showed broader metal ion specificity. They preferred Cd(2+) but were also active with Mn(2+) and Zn(2+), suggesting that binding of the catalytic ion does not require an extremely specific coordination pattern. The unexpected decrease of the catalytic activity of the variants along with the temperature increase suggested that some changes occurred in their structures or the rate-limiting step of the reaction was changed. Two elements of the catalytic core of DNAzyme 1/VIIWS, the nucleotide at position 12 and the three-base-pair hairpin motif, were mutated. The presence of a purine residue at position 12 was crucial for the catalytic activity but the changes at that position had a relatively small influence on the metal ion preferences of this variant. The middle base pair of the three-base-pair hairpin was changed from A-T to C-G interaction. The catalytic activity of the mutated variant was increased with Zn(2+), decreased with Mn(2+), and was not changed in the presence of Cd(2+) ions. Clearly, this base pair was important for defining the metal ion preferences of the DNAzyme 1/VIIWS.


Assuntos
Cádmio/química , DNA Catalítico/química , Sequência de Bases , Catalase/química , Catálise , Domínio Catalítico , Clonagem Molecular , DNA/química , Biblioteca Gênica , Íons , Manganês/química , Metais/química , Dados de Sequência Molecular , Conformação de Ácido Nucleico , Oligonucleotídeos/química , RNA/química , Temperatura , Zinco/química
17.
Biochim Biophys Acta ; 1840(6): 1782-9, 2014 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-24508122

RESUMO

BACKGROUND: Bacitracin is a polypeptide antibiotic active against Gram-positive bacterial strains. Its mechanism of action postulates disturbing the cell wall synthesis by inhibiting dephosphorylation of the lipid carrier. We have discovered that bacitracin induces degradation of nucleic acids, being particularly active against RNA. METHODS: In the examination of the nucleolytic activity of bacitracin several model RNA and DNA oligomers were used. The oligomers were labeled at their 5' ends with (32)P radioisotope and following treatment with bacitracin the cleavage sites and efficiency were determined. RESULTS AND CONCLUSIONS: Bacitracin induces degradation of RNA at guanosine residues, preferentially in single-stranded RNA regions. Bacitracin is also able to degrade DNA to some extent but comparable effects to those observed with RNA require its 10-fold higher concentration. The sites of degradation in DNA are very infrequent and preferentially occur near cytidine residues. Free radicals are not involved in the reaction, and which probably proceeds via a hydrolytic mechanism. The phosphate groups at the cleavage sites are present at the 3' ends of RNA products and at the 5' ends of DNA fragments. Importantly, the presence of EDTA does not influence RNA degradation but completely inhibits the degradation of DNA. For DNA degradation divalent metal ions like Mg(2+), Mn(2+) or Zn(2+) are absolutely necessary. GENERAL SIGNIFICANCE: The ability of bacitracin to degrade nucleic acids via a hydrolytic mechanism was a surprising observation, and it is of interest whether these properties can contribute to its mechanisms of action during antibiotic treatment.


Assuntos
Antibacterianos/farmacologia , Bacitracina/farmacologia , DNA/química , RNA/química , Hidrólise
18.
Inorg Chem ; 52(24): 13927-33, 2013 Dec 16.
Artigo em Inglês | MEDLINE | ID: mdl-24304384

RESUMO

2-Amino-2-hydroxymethyl-propane-1,3-diol, or tris(hydroxymethyl)aminomethane (Tris), is probably the most common biochemical buffer used alone or in combination with other buffers because it is stable, unreactive, and compatible with most proteins and other biomolecules. Being nontoxic, it has even found applications in medicine. Tris is known, however, to coordinate transition metal ions, Cu(II) among them. Although often ignored, this feature affects interactions of Cu(II) ions with biomolecules, as Tris is usually used in high molar excess. Therefore, it is important to have precise knowledge on the stoichiometry, stability, and reactivity of cupric Tris complexes. The literature data are incoherent in this respect. We reinvestigated the complex formation in the Tris-Cu(II) system by potentiometry, UV-vis, ESI-MS, and EPR at a broad range of concentrations and ratios. We found, contrary to several previous papers, that the maximum stoichiometry of Tris to Cu(II) is 2 and at neutral pH, dimeric complexes are formed. The apparent affinity of Tris buffer for Cu(II), determined by the competitivity index (CI) approach [Krezel, A.; Wójcik, J.; Maciejczyk, M.; Bal, W. Chem. Commun. 2003, 6, 704-705] at pH 7.4 varies between 2 × 10(6) and 4 × 10(4) M(-1), depending on the Tris and Cu(II) concentrations and molar ratio.


Assuntos
Complexos de Coordenação/química , Cobre , Modelos Moleculares , Cobre/química , Estabilidade de Medicamentos , Espectrometria de Massas por Ionização por Electrospray , Trometamina
19.
J Inorg Biochem ; 127: 73-8, 2013 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-23880034

RESUMO

Blasticidin S is a representative of the aminoacylnucleoside class of antibiotics and it possesses fungicidal properties against the virulent fungus which causes a serious rice blast disease in Asia. It is widely used to control rice blast by foliar application as a crop protection agent. Serious copper contamination is observed in some areas of China. Moreover, some paddy soils present a potential risk of copper accumulation in the human body through the food chain, leading to several disorders. This work presents the results of combined potentiometric and spectroscopic (UV-visible, EPR, CD, NMR) data which lead to the conclusion that the antibiotic is capable of binding copper, and the resulting complexes are likely to form in the soil. The process of complex formation has a potential influence on the population feeding on a rice-based diet. Moreover, the results of electrophoretic experiments revealed that complexes do not cleave DNA. On the contrary, the presence of blasticidin S may prevent DNA from a Cu(II)-induced damage.


Assuntos
Complexos de Coordenação/química , Cobre/química , Oryza , Complexos de Coordenação/farmacologia , Cobre/metabolismo , DNA/química , Dano ao DNA/efeitos dos fármacos , Espectroscopia de Ressonância de Spin Eletrônica , Humanos , Íons , Modelos Moleculares , Nucleosídeos/química , Nucleosídeos/metabolismo , Nucleosídeos/farmacologia
20.
J Inorg Biochem ; 124: 26-34, 2013 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-23583885

RESUMO

Three representatives of the distinct antibiotics groups: amoxicillin, apramycin and ristomycin A were studied regarding their impact on hepatitis D virus (HDV) ribozyme both in the metal-free form and complexed with copper(II) ions. Hence the Cu(II)-ristomycin A complex has been characterized by means of NMR, EPR, CD and UV-visible spectroscopic techniques and its binding pattern has been compared with the coordination modes estimated previously for Cu(II)-amoxicillin and Cu(II)-apramycin complexes. It has thus been found that all three antibiotics bind the Cu(II) ion in a very similar manner, engaging two nitrogen and two oxygen donors into coordination with the square planar symmetry in physiological conditions. All three tested antibiotics were able to inhibit the HDV ribozyme catalysis. However, in the presence of the complexes, the catalytic reactions were almost completely inhibited. It was important therefore to check whether the complexes used in lower concentrations could inhibit the HDV ribozyme catalytic activity, thus creating opportunities for their practical application. It turned out that the complexes used in the concentrations of 50µM influenced the catalysis much less effectively comparing to the 200 micromolar concentration. The kobs values were lower than those observed in the control reaction, in the absence of potential inhibitors: 2-fold for amoxicillin, ristomycin A and 3.3-fold for apramycin, respectively.


Assuntos
Amoxicilina/química , Cobre/química , Vírus Delta da Hepatite/enzimologia , Nebramicina/análogos & derivados , RNA Catalítico/química , RNA Viral/química , Ristocetina/química , Catálise , Espectroscopia de Ressonância de Spin Eletrônica , Espectroscopia de Ressonância Magnética , Nebramicina/química
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