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De Simone G.,CNR Institute of Biostructure and Bioimaging | Supuran C.T.,University of Florence
Journal of Inorganic Biochemistry | Year: 2012

Carbonic anhydrases (CAs, EC are widespread enzymes in all life kingdoms with five distinct genetic families known to date, the α-, β-, γ-, δ- and ζ-CAs. With the exception of the δ-class, which is less investigated to date, enzymes from the remaining classes found in vertebrates, corals, fungi, bacteria and archaea have been studied for their inhibition with simple inorganic anions as well as more complex inorganic and organic ones. In this paper we review the available data for the inhibition of these enzymes with all anions except sulfonamides and their bioisosteres (sulfamates, sulfamides) which have been reviewed earlier. Anion inhibitors are important both for understanding the inhibition/catalytic mechanisms of these enzymes and for designing novel types of inhibitors which may have clinical applications for the management of a variety of disorders in which CAs are involved. Environmental aspects of CO 2 fixation by CAs present in plants, corals, algae or diatoms and how this may be affected by inhibitors are also discussed. © 2011 Elsevier Inc. All rights reserved.

De Simone G.,CNR Institute of Biostructure and Bioimaging | Supuran C.T.,University of Florence
Biochimica et Biophysica Acta - Proteins and Proteomics | Year: 2010

Isoform IX of the zinc enzyme carbonic anhydrase (CA, EC, CA IX, is a transmembrane protein involved in solid tumor acidification through the HIF-1α activation cascade. CA IX has a very high catalytic activity for the hydration of carbon dioxide to bicarbonate and protons, even at acidic pH values (of around 6.5), typical of solid, hypoxic tumors, which are largely unresponsive to classical chemo- and radiotherapy. Thus, CA IX is used as a marker of tumor hypoxia and as a prognostic factor for many human cancers. CA IX is involved in tumorigenesis through many pathways, such as pH regulation and cell adhesion control. The X-ray structure of the catalytic domain of CA IX has been recently reported, being shown that CA IX has a typical α-CA fold. However, the CA IX structure differs significantly from the other CA isozymes when the protein quaternary structure is considered. Thus, two catalytic domains of CA IX associate to form a dimer, which is stabilized by the formation of an intermolecular disulfide bond. The active site clefts and the proteoglycan (PG) domains are located on one face of the dimer, while the C-termini are located on the opposite face to facilitate protein anchoring to the cell membrane. As all mammalian CAs, CA IX is inhibited by several main classes of inhibitors, such as the inorganic anions, the sulfonamides and their bioisosteres (sulfamates, sulfamides, etc.), the phenols, and the coumarins. The mechanism of inhibition with all these classes of compounds is understood at the molecular level, but the sulfonamides and their congeners have important applications. It has been recently shown that both in vitro, in cell cultures, as well as in animals with transplanted tumors, CA IX inhibition with sulfonamides lead to a return of the extracellular pH to more normal values, which leads to a delay in tumor growth. As a consequence, CA IX represents a promising antitumor target for the development of anticancer agents with an alternative mechanism of action. © 2009 Elsevier B.V. All rights reserved.

Alterio V.,CNR Institute of Biostructure and Bioimaging | Di Fiore A.,CNR Institute of Biostructure and Bioimaging | D'Ambrosio K.,CNR Institute of Biostructure and Bioimaging | Supuran C.T.,University of Florence | De Simone G.,CNR Institute of Biostructure and Bioimaging
Chemical Reviews | Year: 2012

A description of the structural studies on α-Carbonic anhydrases (α-CA) is presented. Main structural features of the catalytically active α-CA isozymes are described and the current state of the art on complexes of hCA II with the principal classes of inhibitors is summarized. Studies have demonstrated that cyanamide acts as a weak inhibitor of the esterase activity of hCA II, interacting with the zinc ion within the enzyme active site. Crystallographic studies have shown that inorganic anions can bind within the CA II active site, either coordinating the catalytic Zn2+ ion or not coordinating it, but being located in its close proximity in a nonmetal site. The effect of the presence of a positive, negative or neutral substituent on benzenesulfonamide CAIs has been investigated to understand the effect of different charges in the CA I and CA II active sites, to discriminate the binding to these two isoforms.

Musumeci D.,University of Naples Federico II | Roviello G.N.,CNR Institute of Biostructure and Bioimaging | Montesarchio D.,University of Naples Federico II
Pharmacology and Therapeutics | Year: 2014

HMGB1 (High-Mobility Group Box-1) is a nuclear protein that acts as an architectural chromatin-binding factor involved in the maintenance of nucleosome structure and regulation of gene transcription. It can be released into the extracellular milieu from immune and non-immune cells in response to various stimuli. Extracellular HMGB1 contributes to the pathogenesis of numerous chronic inflammatory and autoimmune diseases, including sepsis, rheumatoid arthritis, atherosclerosis, chronic kidney disease, systemic lupus erythematosus (SLE), as well as cancer pathogenesis. Interaction of released HMGB1 with the cell-surface receptor for advanced glycation end products (RAGE) is one of the main signaling pathways triggering these diseases. It has been also demonstrated that the inhibition of the HMGB1-RAGE interaction represents a promising approach for the modulation of the inflammatory and tumor-facilitating activity of HMGB1. In this review we describe various approaches recently proposed in the literature to inhibit HMGB1 and the related inflammatory processes, especially focusing on the block of RAGE-HMGB1 signaling. Several strategies are based on molecules which mainly interact with RAGE as competitive antagonists of HMGB1. As an alternative, encouraging results have been obtained with HMGB1-targeting, leading to the identification of compounds that directly bind to HMGB1, ranging from small natural or synthetic molecules, such as glycyrrhizin and gabexate mesilate, to HMGB1-specific antibodies, peptides, proteins as well as bent DNA-based duplexes. Future perspectives are discussed in the light of the overall body of knowledge acquired by a large number of research groups operating in different but related fields. © 2013 Elsevier Inc. All rights reserved.

Improta R.,CNR Institute of Biostructure and Bioimaging | Barone V.,Normal School of Pisa
Angewandte Chemie - International Edition | Year: 2011

The excited states of (dA) 4 oligonucleotides (A=adenine), including the phosphoribose backbone, were studied in water at a fully quantum mechanical level, providing an atomistic description of the main decay paths and a comprehensive interpretation of the experimental data (see picture). After absorption to exciton states delocalized over multiple A bases, the behavior of the excited state is ruled by the interplay of a number of species responsible for different spectral features. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Improta R.,CNR Institute of Biostructure and Bioimaging | Barone V.,Normal School of Pisa
Topics in Current Chemistry | Year: 2015

We review the most significant results obtained in the study of isolated nucleobases in solution by quantum mechanical methods, trying to highlight also the most relevant open issues. We concisely discuss some methodological issues relevant to the study of molecular electronic excited molecular states in condensed phases, focussing on the methods most commonly applied to the study of nucleobases, i.e. continuum models as the Polarizable Continuum Model and explicit solvation models. We analyse how the solvent changes the relative energy of the lowest energy excited states in the Franck–Condon region, their minima and the Conical Intersections among the different states, interpreting the experimental optical spectra, both steady state and time-resolved. Several methods are available for accurately including solvent effects in the Franck–Condon region, and for most of the nucleobases the solvent shift on the different excited states can be considered assessed. The study of the excited state decay, both radiative and non-radiative, in solution still poses instead significant theoretical challenges. © Springer-Verlag Berlin Heidelberg 2014.

Improta R.,CNR Institute of Biostructure and Bioimaging
Journal of Physical Chemistry B | Year: 2012

We here report a fully quantum mechanical study of the main photochemical and photophysical decay routes in aqueous solution of thymine deoxy-dinucleotide (TpT- and TpTNa) and of its analogue locked in C3-endo puckering, characterizing five different representative backbone conformers and discussing the chemical physical effects modulating the yield of the different photoproducts. Our approach is based on time-dependent DFT calculations, using the last generation M052X functional, whereas solvent effects are included by means of the polarizable continuum model. Especially when at least one of the sugars adopts C3-endo puckering, a barrierless path on the bright ππ* excitons leads to the S1/S0 crossing region corresponding to the formation of cyclobutane pyrimidine dimer. Charge transfer excited states involving the transfer of an electron from the 5′ Thy toward the 3′ Thy are involved in the formation of the oxetane intermediate in the path leading to 6-4 pyrimidine pyrimidinone adducts. A non-negligible energy barrier is associated with this latter pathway, which is possible only when one of the two nucleotides adopts C2-endo puckering. Monomer-like decay pathways, involving ππ* or nπ* excited states localized on a single base, are shown to be operative also for loosely stacked bases. © 2012 American Chemical Society.

Improta R.,CNR Institute of Biostructure and Bioimaging
Chemistry - A European Journal | Year: 2014

Herein, a full quantum mechanical study, in solution, of several models of guanine-quadruplex helices, both parallel and antiparallel, containing up to eight guanine residues, in their electronic excited state is reported. By exploiting TD-DFT calculations and including solvent effects by the polarizable continuum model, we provide the first atomistic description of the processes triggered by the absorption of UV light, reproducing and assigning the experimental optical and electronic circular dichroism spectra. The absorbing excited states are delocalized over multiple bases, whereas emission involves a stacked guanine dimer or a monomer. Several states, with a varying degree of localization and charge-transfer character, rule the photoexcited dynamics, which are deeply affected by the quadruplex topology. The lowest excited-state minimum for parallel quadruplex is an asymmetric excimer involving two stacked guanines, with a small charge transfer character, whereas for the anti-parallel structure, with the same topology of the thrombin binding aptamer, it is a fully symmetric excimer, characterized by a strong decrease of the stacking distance. A monomer-like decay path is the most relevant nonradiative decay pathway. Insights on the effect of the ions (K+ or Na+) on the excited state decay are also provided. Shedding light on quadruple helices: Quantum mechanical calculations show what happens when a guanine quadruplex absorbs UV radiation, allowing the assignment of the absorption and fluorescence spectra and giving insights into the factors ruling their photoexcited behavior (see figure). © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Bellia F.,CNR Institute of Biostructure and Bioimaging | Grasso G.,University of Catania
Journal of Mass Spectrometry | Year: 2014

Amylin or islet amyloid polypeptide (IAPP) is a 37-residue peptide hormone secreted from the pancreatic islets into the blood circulation and is cleared by peptidases in the kidney. IAPP aggregates are strongly associated with β-cell degeneration in type 2 diabetes, as demonstrated by the fact that more than 95% of patients exhibit IAPP amyloid upon autopsy. Recently, it has been reported that metal ions such as copper(II) and zinc(II) are implicated in the aggregation of IAPP as well as able to modulate the proteolytic activity of IAPP degrading enzymes. For this reason, in this work, the role of the latter metal ions in the degradation of IAPP by insulin-degrading enzyme (IDE) has been investigated by a chromatographic and mass spectrometric combined method. The latter experimental approach allowed not only to assess the overall metal ion inhibition of the human and murine IAPP degradation by IDE but also to have information on copper- and zinc-induced changes in IAPP aggregation. In addition, IDE cleavage site preferences in the presence of metal ions are rationalized as metal ion-induced changes in substrate accessibility. © 2014 John Wiley & Sons, Ltd.

Improta R.,CNR Institute of Biostructure and Bioimaging | Vitagliano L.,CNR Institute of Biostructure and Bioimaging | Esposito L.,CNR Institute of Biostructure and Bioimaging
Acta Crystallographica Section D: Biological Crystallography | Year: 2015

By combining quantum-mechanical analysis of small model peptides and statistical surveys of high-resolution protein structures, a systematic conformational dependence of bond lengths in polypeptide backbones has been unveiled which involves both the peptide bond (C-O and C-N) and those bonds centred on the Cα atom. All of these bond lengths indeed display a systematic variability in the ψ angle according to both calculations and surveys of protein structures. The overall agreement between the computed and the statistical data suggests that these trends are essentially driven by local effects. The dependence of Cα distances on ψ is governed by interactions between the σ system of the Cα moiety and the C-O π system of the peptide bond. Maximum and minimum values for each bond distance are found for conformations with the specific bond perpendicular and parallel to the adjacent CONH peptide plane, respectively. On the other hand, the variability of the C-O and C-N distances is related to the strength of the interactions between the lone pair of the N atom and the C-O π∗ system, which is modulated by the ψ angle. The C-O and C-N distances are related but their trends are not strictly connected to peptide-bond planarity, although a correlation amongst all of these parameters is expected on the basis of the classical resonance model. © 2015 International Union of Crystallography.

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