1,720,966 research outputs found
Synthesis of labeled peptides and study by EPR methods of their interaction with biological membranes
Full text linkAntimicrobial peptides (AMPs) have been proposed as antitumor molecules since they sometimes displayed selective cytotoxicity towards cancer cells; they have the potential to bypass multi-drug resistance, as they exert their bioactivity by interacting with the membranes, without a specific target. Trichogin GA IV (TG) belongs to lipopeptaibols, a class of antimicrobial peptides characterized by the presence of tetrasubstituted alpha-amino acids in their sequence. Differently from other AMPs, lipopeptaibols present a significant resistance to enzyme hydrolysis. The purpose of the present Ph.D. thesis is the study of the peptide-membrane interaction in healthy and cancer cells as the membrane perturbing ability of these peptides is most likely at the basis of the selective cytotoxicity of the peptides. Working in physiological condition was of primary importance to untangle the specificity of the different peptides for the various cell lines: the membrane properties at physiological temperature are indeed among the main determinants of the peptide-membrane interaction. The study has been conducted using several techniques. Structural features of TG analogs have been quantified to rationally design new analogs with different characteristics. The results of cytotoxicity assays on cancer and healthy cells showed that it is possible to tune the toxicity of TG towards cells with different membrane composition. Fluorescence microscopy showed peptides localization on cell membranes. Electron Paramagnetic Resonance (EPR) spectroscopy is a versatile technique that has been widely exploited to determine the affinity of spin labeled peptides for model and cell membranes and their orientation in the phospholipid bilayer. The obtained results are promising, as we could relate structural modification with variations in TG analogs toxicity, an encourage further studies
Accelerated aging in perinatally HIV-infected children: clinical manifestations and pathogenetic mechanisms
Full text linkBACKGROUND:
Premature aging and related diseases have been documented in HIV-infected adults. Data are now emerging also regarding accelerated aging process in HIV-infected children.
METHODS:
A narrative review was performed searching studies on PubMed published in English language in 2004-2017, using appropriate key words, including "aging", "children", "HIV", "AIDS", "immunosenescence", "pathogenesis", "clinical conditions".
RESULTS:
Premature immunosenescence phenotype of B and T cells in HIV-infected children is mediated through immune system activation and chronic inflammation. Ongoing inflammation processes have been documented by increased levels of pathogen-associated molecular patterns (PAMPS), increased mitochondrial damage, higher levels of pro-inflammatory cytokines, and a positive correlation between sCD14 levels and percentages of activated CD8+ cells. Other reported features of premature aging include cellular replicative senescence, linked to an accelerated telomeres shortening. Finally, acceleration of age-associated methylation pattern and other epigenetic modifications have been described in HIV-infected children. All these features may favor the clinical manifestations related to premature aging. Lipid and bone metabolism, cancers, cardiovascular, renal, and neurological systems should be carefully monitored, particularly in children with detectable viremia and/or with CD4/CD8 ratio inversion.
CONCLUSION:
Aging processes in children with HIV infection impact their quality and length of life. Further studies regarding the mechanisms involved in premature aging are needed to search for potential targets of treatment
Interaction of hydrophobic and amphipathic antimicrobial peptides with lipid bicelles
No full textBicelles are model membrane systems that can be macroscopically oriented in a magnetic field at physiological temperature. The macroscopic orientation of bicelles allows to detect, by means of magnetic resonance spectroscopies, small changes in the order of the bilayer caused by solutes interacting with the membrane. These changes would be hardly detectable in isotropic systems such as vesicles or micelles. The aim of this work is to show that bicelles represent a convenient tool to investigate the behavior of antimicrobial peptides (AMPs) interacting with membranes, using electron paramagnetic resonance (EPR) spectroscopy. We performed the EPR experiments on spin-labeled bicelles using various AMPs of different length, charge, and amphipathicity: alamethicin, trichogin GA IV, magainin 2, HP(2–20), and HPA3. We evaluated the changes in the order parameter of the spin-labeled lipids as a function of the peptide-to-lipid ratio. We show that bicelles labeled at position 5 of the lipid chains are very sensitive to the perturbation induced by the AMPs even at low peptide concentrations. Our study indicates that peptides that are known to disrupt the membrane by different mechanisms (i.e., alamethicin vs magainin 2) show very distinct trends of the order parameter as a function of peptide concentration. Therefore, spin-labeled bicelles proved to be a good system to evaluate the membrane disruption mechanism of new AMPs
Lipid Bicelles As A Model System To Study the Interaction of Hydrophobic and Amphipathic Antimicrobial Peptides With Lipid Membranes
No full text
Liponitroxides: EPR study and their efficacy as antioxidants in lipid membranes
No full textA series of lipid-functionalized nitroxides having a pyrroline nitroxide moiety linked either to a glycerol or to
a steroid unit has been synthesized, and their inclusion inside phospholipid bilayers has been investigated by
Electron Paramagnetic Resonance (EPR) spectroscopy. The antioxidant behavior of these nitroxides has
been studied in azo-initiator induced lipid peroxidation by means of the Thiobarbituric Acid Reactive
Species (TBARS) assay; a correlation with their penetration depth within the bilayer has been found. The
possible mechanisms involved in the antioxidant action have been considered, discussed and alternative
pathways have been suggested for the synthesized liponitroxides due to their different localization. The
steroid derivative is limited to scavenging radicals that are generated in the aqueous phase, while the
glycerolipids can also act as chain breaking antioxidants
The rational search for selective anticancer derivatives of the peptide Trichogin GA IV: a multi-technique biophysical approach
Full text linkPeptaibols are peculiar peptides produced by fungi as weapons against
other microorganisms. Previous studies showed that peptaibols are
promising peptide-based drugs because they act against cell membranes
rather than a specific target, thus lowering the possibility of the
onset of multi-drug resistance, and they possess non-coded alpha-amino
acid residues that confer proteolytic resistance. Trichogin GA IV (TG)
is a short peptaibol displaying antimicrobial and cytotoxic activity. In
the present work, we studied thirteen TG analogues, adopting a
multidisciplinary approach. We showed that the cytotoxicity is tuneable
by single amino-acids substitutions. Many analogues maintain the same
level of non-selective cytotoxicity of TG and three analogues are
completely non-toxic. Two promising lead compounds, characterized by the
introduction of a positively charged unnatural amino-acid in the
hydrophobic face of the helix, selectively kill T67 cancer cells without
affecting healthy cells. To explain the determinants of the
cytotoxicity, we investigated the structural parameters of the peptides,
their cell-binding properties, cell localization, and dynamics in the
membrane, as well as the cell membrane composition. We show that, while
cytotoxicity is governed by the fine balance between the amphipathicity
and hydrophobicity, the selectivity depends also on the expression of
negatively charged phospholipids on the cell surface
Insights into Peptide-Membrane Interactions of Newly Synthesized, Nitroxide-Containing Analogs of the Peptaibiotic Trichogin GA IV Using EPR
Full text linkTrichogin GA IV is a short-length (10-amino acid long), mostly hydrophobic peptaibiotic with an N-terminal fatty acyl chain and a C-terminal 1,2-amino alcohol. A cardinal role of the terminal moieties in the cytotoxic activity of trichogin has been recently found. Previously, peptide orientation and dynamics of trichogin analogs in the membrane were studied using methyl ester derivatives. Therefore, in the present work we synthesized several trichogin analogs with naturally occurring terminal groups to verify whether these moieties have any effect on peptide-membrane interaction. These trichogin analogs, both neutral and carrying a side-chain, positively charged Lys residue, bear the nitroxide-containing α-amino acid TOAC to study them using EPR spectroscopy. Vesicles were used to investigate orientation and penetration depth of the peptide at room temperature. Bicelles were employed to evaluate the order, dynamics, and orientation of the peptide at a near physiological temperature. In addition, the position of the N-terminal 1-octanoyl chain in the membrane was studied by labeling it with a nitroxide. The secondary structure of the peptides in vesicles was studied by CD spectroscopy showing that they adopt a mostly α-helical structure. In vesicles, the analogs insert below the lipid headgroups with the helix axis oriented parallel to the membrane surface at a peptide-to-lipid (P:L) ratio of 1:100. The presence of the single, positively charged Lys residue does not alter the orientation adopted by the peptides. In bicelles at P:L ratios 1:100 and 1:60, the peptide adopts a transmembrane orientation characterized by a very low orientational order, whereas at the 1:15 P:L ratio it severely disrupts the membrane. Our data shows that overall orientation and insertion in model membranes of the native trichogin GA IV are strictly comparable to those of its methyl ester analogs previously examined. This article is protected by copyright. All rights reserved
Going Beyond Counting First Authors in Author Co-citation Analysis
Full text linkThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
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