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Francesca DE RIENZO

Personale tecnico amministrativo
Dipartimento di Scienze Chimiche e Geologiche - Sede Dipartimento di Scienze Chimiche e Geologiche

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2014 - Synthesis and structureeactivity relationship studies in serotonin 5-HT4 receptor ligands based on a benzo[de][2,6]naphthridine scaffold [Articolo su rivista]
Federica, Castriconi; Marco, Paolino; Germano, Giuliani; Maurizio, Anzini; Giuseppe, Campiani; Laura, Mennuni; Chiara, Sabatini; Marco, Lanza; Gianfranco, Caselli; DE RIENZO, Francesca; Menziani, Maria Cristina; Maria, Sbraccia; Paola, Molinari; Tommaso, Costa; Andrea, Cappelli

A small series of serotonin 5-HT4 receptor ligands has been designed from flexible 2-methoxyquinoline compounds 7a,b by applying the conformational constraint approach. Ligands 7a,b and the corresponding conformationally constrained analogues 8aeg were synthesized and their interactions with the 5-HT4 receptor were examined by measuring both binding affinity and the ability to promote or inhibit receptoreG protein coupling. Ester derivative 7a and conformationally constrained compound 8b were demonstrated to be the most interesting compounds showing a nanomolar 5-HT4R affinity similar to that shown by reference ligands cisapride (1) and RS-23,597-190 (4). The result was rationalized by docking studies in term of high similarity in the binding modalities of flexible 7a and conformationally constrained 8b. The intrinsic efficacy of some selected ligands was determined by evaluating the receptoreG protein coupling and the results obtained demonstrated that the nature and the position of substituents play a critical role in the interaction of these ligands with their receptor.

2013 - Approaching the 5-HT3 receptor heterogeneity by computational studies of the transmembrane and intracellular domains [Articolo su rivista]
Marta Del, Cadia; DE RIENZO, Francesca; Menziani, Maria Cristina

5-hydroxytryptamine type-3 receptor (5-HT3), an important target of many neuroactive drugs, is a cation selective transmembrane pentamer whose functional stoichiometries and subunit arrangements are still debated, due to the extreme complexity of the system. The three dimensional structure of the 5-HT3R subunits has not been solved so far. Moreover, most of the available structural and functional data is related to the extracellular ligand-binding domain, whereas the transmembrane and the intracellular receptor domains are far less characterised, although they are crucial for receptor function. Here, for the first time, 3D homology models of the transmembrane and the intracellular receptor domains of all the known human 5-HT3 subunits have been built and assembled into homopentameric (5-HT3AR, 5-HT3BR, 5-HT3CR, 5-HT3DR and 5-HT3ER) and heteropentameric receptors (5-HT3AB, 5-HT3AC, 5-HT3AD and 5-HT3AE), on the basis of the known three-dimensional structures of the nicotinic-acetylcholine receptor and of the ligand gated channel from Erwinia chrysanthemi. The comparative analyses of sequences, modelled structures, and computed electrostatic properties of the single subunits and of the assembled pentamers shed new light both on the stoichiometric composition and on the physicochemical requirements of the functional receptors. In particular, it emerges that a favourable environment for the crossing of the pore at the transmembrane and intracellular C terminus domain levels by Ca2+ ions is granted by the maximum presence of two B subunits in the 5-HT3 pentamer.

2013 - Exploring a potential palonosetron allosteric binding site in the 5-HT 3 receptor [Articolo su rivista]
Del Cadia, M; DE RIENZO, Francesca; Weston D., A; Thompson A., J; Menziani, Maria Cristina; Lummis, S. C. R.

Palonosetron (Aloxi) is a potent second generation 5-HT3 receptor antagonist whose mechanism of action is not yet fully understood. Palonosetron acts at the 5-HT3 receptor binding site but recent computational studies indicated other possible sites of action in the extracellular domain. To test this hypothesis we mutated a series of residues in the 5-HT3A receptor subunit (Tyr73, Phe130, Ser 163, and Asp165) and in the 5-HT3B receptor subunit (His73, Phe130, Glu170, and Tyr143) that were previously predicted by in silico docking studies to interact with palonosetron. Homomeric (5-HT3A) and heteromeric (5-HT3AB) receptors were then expressed in HEK293 cells to determine the potency of palonosetron using both fluorimetric and radioligand methods to test function and ligand binding, respectively. The data show that the substitutions have little or no effect on palonosetron inhibition of 5-HT-evoked responses or binding. In contrast, substitutions in the orthosteric binding site abolish palonosetron binding. Overall, the data support a binding site for palonosetron at the classic orthosteric binding pocket between two 5-HT3A receptor subunits but not at allosteric sites previously identified by in silico modelling and docking.

2012 - A first step towards the understanding of the 5-HT3 receptor subunitheterogeneity from a computational point of view [Articolo su rivista]
DE RIENZO, Francesca; DEL CADIA, Marta; Menziani, Maria Cristina

The functional serotonin type-3 receptor (5-HT3-R), which is the target of many neuroactive drugs, isknown to be a homopentamer made of five identical subunits A (5-HT3A-R) or a binary heteropentamermade of subunits A and B (5-HT3A/B-R) with a still debated arrangement and stoichiometry. Thiscomplex picture has been recently further complicated by the discovery of additional 5-HT3-R subunits,C, D, and E, which, similarly to the B subunit, are apparently able to form functional receptors only ifco-expressed with subunit A. Being the binding site for both serotonin and antagonists (i.e. drugs)located at the extracellular interface between two adjacent subunits, the large variability of the 5-HT3-Rcomposition becomes a crucial issue, since it can originate many different interfaces providing nonequivalentligand binding sites and complicating the pharmacological modulation. Here, the different5-HT3-R interfaces are analysed, on the bases of the structural conformations of previously built 3Dhomology models and of the known subunit sequences, by addressing their physicochemicalcharacterization. The results confirm the presence of an aromatic cluster located in the core of the A–Ainterface as a key determinant for having an interface both stable and functional. This is used as adiscriminant to make hypotheses about the capability of all the other possible interfaces constituted bythe known 5-HT3-R sequences A, B, C, D, and E to build active receptors.

2012 - Computational Insights into ADAMTS4, ADAMTS5 andMMP13 Inhibitor Selectivity [Articolo su rivista]
FILOMIA, Federico; SAXENA, PUNEET; DURANTE, Caterina; DE RIENZO, Francesca; COCCHI, Marina; MENZIANI, Maria Cristina

The results obtained by means of Molecular Dynamicssimulations and Multiway Explorative Data Analysison ADAMTS4, ADAMTS5 and MMP13 complexed with Marimastatand two cis-1(S)2(R)-amino-2-indanol ligands suggestthat determinant characteristics for ligand binding andselectivity among the three enzymes are to be found in thedifferent protein conformation flexibility. Moreover, the role of the TS-domain in the inhibitor binding to ADAMTS enzymeshas been investigated for the first time in this work.The results obtained suggest that the influence of the TSdomainon the S1’ loop fluctuations of ADAMTS4 andADAMTS5 could be exploited for the design of therapeuticsfor chronic osteoarthritis diseases.

2012 - The extracellular subunit interface of the 5-HT3 Receptors: a Computational Alanine Scanning Mutagenesis study Journal of Biomolecular Structure and Dynamics [Articolo su rivista]
DE RIENZO, Francesca; MOURA BARBOSA, ARMENIO JORGE; M. A. S., Perez; P. A., Fernandes; M. J., Ramos; Menziani, Maria Cristina

The functional serotonin 5-HT type-3 (5-HT3) receptor, the target of many neuroactive drugs, is known to be apseudo-symmetric pentamer made either of five identical subunits A (homomeric 5-HT3A-R) or of subunits A and B(heteromeric 5-HT3A/B-R) in a still debated arrangement. The serotonin binding site is located in the extracellular region,at the interface between two monomers, called the principal and the complementary subunits. The results of moleculardynamics simulations and computational alanine scanning mutagenesis studies applied here to the homomeric human5-HT3A-R disclose an aromatic “hot” cluster in the centre of the interface formed by residues W178 (principal subunit),Y68, Y83, W85 and Y148 (complementary subunit). Moreover, investigation of the coupling of agonist/antagonist bindingto channel activation/inactivation points out the presence of two putative functional pathways at the subunit interface:W116-H180-L179-W178-E124-F125 (principal subunit) and Y136-Y138-Y148-W85-(P150) (complementary subunit),where W178 and Y148 appear to be critical residues for the binding/activation mechanism. Finally, direct comparison ofthe main features shown by the AA interface in the human 5-HT3A-R with those of the BB interface in the homopentamerichuman 5-HT3B-R provides interesting clues about the possible reasons that cause the 5-HT3B-R not to befunctional.

2011 - Bivalent Ligands for the Serotonin 5-HT3 Receptor [Articolo su rivista]
A., Cappelli; M., Manini; M., Paolino; A., Gallelli; M., Anzini; L., Mennuni; DEL CADIA, Marta; DE RIENZO, Francesca; Menziani, Maria Cristina; S., Vomero

The serotonin 5-HT3 receptor is a ligand-gated ion channel, which by virtue of its pentameric architecture, canbe considered to be an intriguing example of intrinsicallymultivalent biological receptors. This paper describes a generaldesign approach to the study of multivalency in this multimericion channel. Bivalent ligands for 5-HT3 receptor have beendesigned by linking an arylpiperazine moiety to probes showing different functional features. Both homobivalent and hetero- bivalent ligands have shown 5-HT3 receptor affinity in the nanomolar range, providing evidence for the viability of our design approach. Moreover, the high affinity shown by homo- bivalent ligands suggests that bivalency is a promising approachin 5-HT3 receptor modulation and provides the rational basis for applying the concepts of multivalency to the study of 5-HT3 receptor function.

2010 - Computational analysis of ligand recognition sites of homo- andheteropentameric 5-HT3 receptors [Articolo su rivista]
Moura Barbosa, Armenio Jorge; De Rienzo, Francesca; M. J., Ramos; Menziani, Maria Cristina

Inhibition of the 5-hydroxytryptamine receptor (5-HT3R), a member of the Cys-loop superfamily ofLigand-Gated Ion Channels (LGICs), has been recognized to have important antiemetic effects. Withrespect to the many other drugs already in use, such as the first generation 5-HT3R antagonist granisetron,palonosetron, a second generation antagonist, clearly demonstrates superior inhibition potencytowards the 5-HT3Rs. Five different receptor monomers, the 5-HT3R AeE, have been identified althoughthe A and B subunits are the only known to build functional receptors, the homopentameric 5-HT3AR andthe heteropentameric 5-HT3BeAR (with BBABA subunit arrangement). At present, however, no threedimensionalstructure has been reported for any of the 5-HT3R subunits. To understand the bindingproperties of agonists and antagonists, models of the extracellular portion of the 5-HT3R A and B subunitsare built and assembled into the receptor (homo- and hetero-) pentameric structure on the basis of theknown three-dimensional structure of the nicotinic-acetylcholine receptor (nACh-R). The results ofdocking studies of the natural agonist serotonin and the antagonists palonosetron and granisetron intothe modelled homomeric and heteromeric 5-HT3R binding interfaces, provide a possible rationalizationboth of the higher potency of palonosetron with respect to other antagonists, and of its previouslyreported allosteric binding and positive cooperativity properties

2010 - Disruption of the H-bond network in the main access channel of catalase–peroxidasemodulates enthalpy and entropy of Fe(III) reduction [Articolo su rivista]
J., Vlasits; Bellei, Marzia; C., Jakopitsch; De Rienzo, Francesca; P. G., Furtmüller; M., Zamocky; Sola, Marco; Battistuzzi, Gianantonio; C., Obinger

Catalase–peroxidases are the only heme peroxidases with substantial hydrogen peroxide dismutation 28activity. In order to understand the role of the redox chemistry in their bifunctional activity, catalatically- 29active and inactive mutant proteins have been probed in spectroelectrochemical experiments. In detail, wild- 30type KatG from Synechocystis has been compared with variants with (i) disrupted KatG-typical adduct 31(Trp122-Tyr249-Met275), (ii) mutation of the catalytic distal His123-Arg119 pair, and (iii) altered 32accessibility to the heme cavity (Asp152, Ser335) and modified charge at the substrate channel entrance 33(Glu253). A valuable insight into the mechanism of reduction potential (E°′) modulation in KatG has been 34obtained from the parameterization of the corresponding enthalpic and entropic components, determined 35from the analysis of the temperature dependence of E°′. Moreover, model structures of ferric and ferrous 36Synechocystis KatG have been computed and used as reference to analyze and discuss the experimental data. 37The results, discussed by reference to published resonance Raman data on the strength of the proximal iron- 38imidazole bond and catalytic properties, demonstrate that E°′ of the Fe(III)/Fe(II) couple is not strongly 39correlated with the bifunctional activity. Besides the importance of an intact Trp-Tyr-Met adduct, it is the 40architecture of the long and constricted main channel that distinguishes KatGs from monofunctional 41peroxidases. An ordered matrix of oriented water dipoles is important for H2O2 oxidation. Its disruption 42results in modification of enthalpic and entropic contributions to E°′ that reflect reduction-induced changes 43in polarity, electrostatics, continuity and accessibility of solvent to the metal center as well as alterations in 44solvent reorganization.

2010 - Insight into MAPK P38α DFG-Flip mechanism by accelerated molecular dynamics [Articolo su rivista]
Filomia, Federico; DE RIENZO, Francesca; Menziani, Maria Cristina

The DFG motif at the beginning of the activation loop of the MAPK p38a undergoes a local structural reorganizationupon binding of allosteric type-II and type-III inhibitors, which causes the residue F169 to movefrom a buried conformation (defined as DFG-in) to a solvent exposed conformation (defined as DFG-out).Although both experimental and computer simulation studies had been performed with the aim ofunveiling the details of the DFG-in to DFG-out transition, the molecular mechanism is still far from beingunequivocally depicted.Here, the accelerated molecular dynamics (AMD) technique has been applied to model the active loopflexibility of p38a and sample special protein conformations which can be accessible only in some conditionsor time periods. Starting from the assumption of an experimentally known initial and final state ofthe protein, the study allowed the description of the interaction network and the structural intermediateswhich lead the protein to change its loop conformation and active site accessibility. Besides a few importanthydrogen bond interactions, a primary role seems to be played by cation–p interactions, involvingthe DFG-loop residue F169, which participate in the stabilization of an intermediate conformation andin its consequent transition to the DFG-out conformation. From this study, insights which may prove usefulfor inhibitor design and/or site directed mutagenesis studies are derived.

2010 - Protein-surface interactions: challenging experiments and computations [Articolo su rivista]
O., Cohavi; S., Corni; DE RIENZO, Francesca; DI FELICE, Rosa; K. E., Gottschalk; M., Hoefling; D., Kokh; Molinari, Elisa; G., Schreiber; A., Vaskevich; R. C., Wade

Protein-surface interactions are fundamental in natural processes, and have great potential for applications ranging from nanotechnology to medicine. A recent workshop highlighted the current achievements and the main challenges in the field. Copyright (C) 2009 John Wiley & Sons, Ltd.

2009 - Progress Towards the Identification of New Aggrecanase Inhibitors [Articolo su rivista]
DE RIENZO, Francesca; Saxena, Puneet; Filomia, Federico; G., Caselli; F., Colace; L., Stasi; A., Giordani; Menziani, Maria Cristina

Degenerative diseases are still a challenging issue in clinical therapy; even though in several cases it is possible to treat symptoms, drugs able to block disease progression are lacking at present. Osteoarthritis (OA) and Rheumatoid Arthritis (RA) are degenerative diseases leading to serious cartilage destruction, affecting joint functions and giving rise to restricted movement, pain and chronic disability. Current clinical treatment for arthritis is confined to Non Steroidal Anti-Inflammatory Drugs (NSAIDs), which are effective in treating symptoms but fail to block the progression of the disease. Matrix Metalloproteases (MMPs) inhibitors have been clinically studied as possible drugs for cartilage degradation prevention. However, their clinical use has been limited by severe side-effects. Aggrecan, which plays a fundamental role in maintaining the structural and mechanical properties of cartilage, has recently been found to be specifically cleaved by "aggrecanases". Aggrecanases are multidomain zinc metalloproteases, different from MMPs, which cleave the aggrecan within the interglobular domain (IGD). Aggrecan breakdown at this site has been found to be crucial for cartilage degradation. These new findings re-addressed the interest of the research for new arthritis therapeutic agents focusing on aggrecanases rather than on MMPs. This review is meant to provide a critical appraisal of the ongoing developments of Zn-chelating and non chelating aggrecanase inhibitors, with a particular emphasis on the related structure-activity relationships (SARs), in the light of the protein structural information recently made available.

2008 - Cloning, expression and physico-chemical characterization of a di-heme cytochrome c4 from the psychrophilic bacterium Pseudoalteromonas haloplanktis TAC 125 [Articolo su rivista]
DI ROCCO, Giulia; Battistuzzi, Gianantonio; Borsari, Marco; DE RIENZO, Francesca; Ranieri, Antonio; M. L., Tutino; Sola, Marco

The 20-kDa di-heme cytochrome c4 from thepsycrophilic bacterium Pseudoalteromonas haloplanktisTAC 125 was cloned and expressed in Escherichia coli andinvestigated through UV–vis and 1H NMR spectroscopiesand protein voltammetry. The model structure was computedusing the X-ray structure of Pseudomonas stutzericytochrome c4 as a template. The protein shows unprecedentedproperties within the cytochrome c4 family,including (1) an almost nonpolar surface charge distribution,(2) the absence of high-spin heme Fe(III) states,indicative of a thermodynamically stable and kineticallyinert axial heme His,Met coordination, and (3) identical E0values for the two heme centers (+0.322 V vs the standardhydrogen elecrode). At pH extremes, both heme groupsundergo the ‘‘acid’’ and ‘‘alkaline’’ conformational transitionstypical of class I cytochromes c, involving ligandexchangeequilibria, whereas at intermediate pH valuestheir electronic properties are sensitive to several residueionizations.

2007 - Free energy of transition for the individual alkaline conformers of yeast iso-1-cytochrome c [Articolo su rivista]
Battistuzzi, Gianantonio; Borsari, Marco; DE RIENZO, Francesca; DI ROCCO, Giulia; Ranieri, Antonio; Sola, Marco

Direct protein electrochemistry was used to obtain the thermodynamic parameters of transition from the native (state III) to the alkaline (state IV) conformer for untrimethylated Saccharomyces cerevisiae iso-1-cytochrome c expressed in E. coli and its single and multiple lysine-depleted variants. In these variants, one or more of the lysine residues involved in axial Met substitution (Lys72, Lys73, and Lys79) was mutated to alanine. The aim of this work is to determine the thermodynamic affinity of each of the substituting lysines for the heme iron and evaluate the interplay of enthalpic and entropic factors. The equilibrium constants for the deprotonation reaction of Lys72, 73, and 79 were computed for the minimized MD average structures of the wild-type and mutated proteins, applying a modified Tanford-Kirkwood calculation. Solvent accessibility calculations for the substituting lysines in all variants were also performed. The transition enthalpy and entropy values within the protein series show a compensatory behavior, typical of a process involving extensive solvent reorganization effects. The experimental and theoretical data indicate that Lys72 most readily deprotonates and replaces M80 as the axial heme iron ligand, whereas Lys73 and Lys79 show comparably higher pK(a) values and larger transition free energies. A good correlation is found within the series between the lowest calculated Lys pK(a) value and the corresponding experimental pK(a) value, which can be interpreted as indicative of the deprotonating lysine itself acting as the triggering group for the conformational transition. The triple Lys to Ala mutant, in which no lysine residues are available for heme iron binding, features transition thermodynamics consistent with a hydroxide ion replacing the axial methionine ligand.

2006 - A computational protocol to probe the role of solvation effects on the reduction potential of azurin mutants [Articolo su rivista]
V., Barone; DE RIENZO, Francesca; E., Langella; Menziani, Maria Cristina; N., Rega; Sola, Marco

Semiquantitative relationships between thermodynamic parameters of Cu2+ reduction experimentally measured for a series of azurin mutants and the solvation free energy of the oxidized state of the proteins were derived. Solvation free energy calculations were carried out within an ONIOM/PCM scheme specifically adapted to this protein series. The method proved to be able to capture the main determinants of the measured reduction parameters, providing satisfactory predictions of the E degrees'.

2006 - Redox properties of the Fe3+/Fe2+ couple in Arthromyces ramosus class II peroxidase and its cyanide adduct [Articolo su rivista]
Battistuzzi, Gianantonio; Bellei, Marzia; DE RIENZO, Francesca; Sola, Marco

The thermodynamics of the one-electron reduction of the ferric heme in free and cyanide-bound Arthromyces ramosus peroxidase (ARP), a class II plant peroxidase, were determined through spectro-electrochemical experiments. The data were compared with those for class III horseradish peroxidase C (HRP) and its cyanide adduct, and were interpreted in terms of ligand binding features, electrostatic effects and solvent accessible surface area of the heme group and of catalytically relevant residues in the heme distal site. The E-o' values for free and cyanide-bound ARP (-0.183 and -0.390 V, respectively, at 25 degrees C and pH 7) are higher than those for HRP and HRP-CN. ARP features an enthalpic stabilization of the ferrous state and a remarkably negative reduction entropy, which are both unprecedented for heme peroxidases. Once the compensatory contributions of solvent reorganization are partitioned from the measured reduction enthalpy, the resulting protein-based Delta H-rc(int)degrees' value for ARP turns out to be less positive than that for HRP by + 10 kJ mol(-1). The smaller stabilization of the oxidized heme in ARP most probably results from the less pronounced anionic character of the proximal histidine, and the decreased polarity in the heme distal site as compared with HRP, as indicated by the X-ray structures. The surprisingly negative Delta S-rc degrees' value for ARP is the result of peculiar reduction-induced solvent reorganization effects.

2005 - Role of the electronic properties of azurin active site in the electron-transfer process [Articolo su rivista]
Corni, S; DE RIENZO, Francesca; DI FELICE, Rosa; Molinari, Elisa

Electron transfer proteins, such as azurin (a blue copper protein), are promising candidates for the implementation of biomolecular nanoelectronic devices. To understand the details of electron transfer in redox active azurin molecules, we performed plane-wave pseudo-potential density functional theory (DFT) calculations of the protein active site in the two possible oxidation states Cu(I) and Cu(II). The ab initio results are used to discuss how the electronic spectrum and wavefunctions may mediate the shuttling of electrons through the copper ion. We find that the Cu-ligand hybridization is very similar in the two charge states of the metal center, but the energy spectrum changes substantially. This result might indicate important effects of electronic correlations in the redox activity and consequent electron transfer through the Cu site.

2004 - Computational approaches to structural and functional analysis of plastocyanin and other blue copper proteins [Articolo su rivista]
DE RIENZO, Francesca; Rr, Gabdoulline; Rc, Wade; Sola, Marco; Menziani, Maria Cristina

Computational techniques are becoming increasingly important in structural and functional biology, in particular as tools to aid the interpretation of experimental results and the design of new systems. This review reports on recent studies employing a variety of computational approaches to unravel the microscopic details of the structure-function relationships in plastocyanin and other proteins belonging to the blue copper superfamily. Aspects covered include protein recognition, electron transfer and protein-solvent interaction properties of the blue copper protein family. The relevance of integrating diverse computational approaches to address the analysis of a complex protein system, such as a cupredoxin metalloprotein, is emphasized.

2003 - Development of an IL-6 antagonist peptide that induces apoptosis in IL-6 dependent 7TD1 cells. [Articolo su rivista]
Manfredini, Rossella; Tenedini, Elena; M., Siena; Tagliafico, Enrico; Montanari, Monica; Grande, Alexis; ZANOCCO MARANI, Tommaso; C., Poligani; Zini, Roberta; A., Bergamaschi; DE RIENZO, Francesca; DE BENEDETTI, Pier Giuseppe; Menziani, Maria Cristina; Ferrari, Sergio

Interleukin-6 (IL-6) is a pleiotropic cytokine involved in the regulation of proliferation and differentiation of hematopoietic cells and in the pathogenesis of many diseases, including multiple myeloma. This study pursues a way to interfere with IL-6 pathway in an attempt to modulate its biological activity. Here we describe the rational design and biological evaluation of peptides able to antagonize the murine IL-6 activity by interfering with IL-6 Receptor alpha in 7TD1 cells, a IL-6-dependent B-cell line. Of the peptide tested, only Guess 4a is capable of interfering with IL-6 transducing pathway, therefore inducing growth arrest and apoptosis of 7TD1 cells.

2003 - Electronic rectification in protein devices [Articolo su rivista]
Rinaldi, R; Biasco, A; Maruccio, G; Arima, V; Visconti, P; Cingolani, R; Facci, P; DE RIENZO, Francesca; DI FELICE, Rosa; Molinari, Elisa; Verbeet, Mp; Canters, Gw

We show that the electron-transfer protein azurin can be used to fabricate biomolecular rectifiers exploiting its native redox properties, chemisorption capability and electrostatic features. The devices consist of a protein layer interconnecting nanoscale electrodes fabricated by electron beam lithography. They exhibit a rectification ratio as large as 500 at 10 V, and operate at room temperature and in air.

2002 - Biologically relevant properties of copper-containing proteins analysed by means of semi-quantitative and quantitative theoretical descriptors [Capitolo/Saggio]
DE RIENZO, Francesca; R. R., Gabdoulline; R. C., Wade; Menziani, Maria Cristina

Biologically relevant properties of copper-containing proteins analysed by means of semi-quantitative and quantitative theoretical descriptors

2002 - Solid-State Molecular Rectifier Based on Self-Organized Metalloproteins [Articolo su rivista]
R., Rinaldi; A., Biasco; G., Maruccio; R., Cingolani; D., Alliata; L., Andolfi; P., Facci; DE RIENZO, Francesca; DI FELICE, Rosa; Molinari, Elisa

A protein-based solid-state electronic device that operates in air is reported; it is fabricated by interconnecting an azurin monolayer immobilized on SiO2 with two gold nanoelectrodes. The operating mechanism is ascribed to resonant tunneling through the redox active center, combined with macroscopic polarization of the oriented self-organized protein layer. The Figure shows the molecular electrostatic potential of the oxidized protein.

2002 - Theoretical descriptors for the quantitative rationalisation of plastocyanin mutant functional propertiess [Articolo su rivista]
DE RIENZO, Francesca; G., Grant; Menziani, Maria Cristina

A quantitative rationalisation of the effect of specific amino acids on the recognition process and redox characteristics of plastocyanin towards cytochrome f, as determined by point mutation experiments, has been attempted in this study. To achieve this goal we derived theoretical descriptors directly from the three-dimensional structure of the plastocyanin mutants, in the same manner as it is usually done for small drug-like molecules. The protein descriptors computed can be related to: (a) the electrostatic and dipole-dipole interactions, effective at long distance; (b) polar interactions whose features are encoded by charged partial surface area descriptors; (c) the propensity of the surface residues to form hydrogen bonding interactions; and (d) dispersion and repulsive interactions. Moreover, an estimation of mutation-dependent variation of redox potential observed has been obtained by electrostatic free energy calculations. The quantitative structure-activity relationship (QSAR) models offer structural interpretation of the point mutation experiment responses and can be of help in the design of new protein engineering experiments.

2001 - A computational model of the 5-HT3 receptor extracellular domain: search for ligand binding sites [Articolo su rivista]
Menziani, Maria Cristina; DE RIENZO, Francesca; A., Cappelli; M., Anzini; DE BENEDETTI, Pier Giuseppe

A three-dimensional model of the 5-HT3 receptor extracellular domain has been derived on the basis of the nicotinic acetylcholine receptor model recently published by Tsigelny et al. Maximum complementarity between the position and characteristics of mutated residues putatively involved in ligand interaction and the pharmacophoric elements derived by the indirect approach applied on several series of 5-HT3 ligands have been exploited to gain insights into the ligand binding modalities and to speculate on the mechanistic role of the structural components. The analysis of the three-dimensional model allows one to distinguish among amino acids that exert key roles in ligand interactions, subunit architecture, receptor assembly and receptor dynamics. For some of these, alternative roles with respect to the ones hypothesized by experimentalists are assigned. Different binding modalities for agonists and antagonists are highlighted, and residues which probably play a role in the transduction of binding into a change in conformational state of the receptor are suggested.

2001 - Control of metalloprotein reduction potential: The role of electrostatic and solvation effects probed on plastocyanin mutants [Articolo su rivista]
Battistuzzi, Gianantonio; Borsari, Marco; Loschi, Lodovica; Menziani, Maria Cristina; DE RIENZO, Francesca; Sola, Marco

The changes in the thermodynamics of Cu(II) reduction for spinach plastocyanin induced by point mutations altering the electrostatic potential in proximity of the copper center were determined through variable temperature direct electrochemistry experiments. In particular, the functionally important surface residues Leu12 and Gln88 were replaced with charged and polar residues, and Asn38 was substituted with Asp. The mutational variations of the reduction enthalpy and entropy were analyzed with a QSPR (quantitative structure-property relationships) approach, employing global and local theoretical descriptors defined and computed on the three-dimensional protein structure. The correlations found are informative on how electrostatic and solvation effects control the E degrees' values in this species through the combined effects on the reduction enthalpy and entropy. The changes in reduction enthalpy can be justified with electrostatic considerations. Most notably, enthalpy-entropy compensation phenomena play a significant role: the entropic effects due to the insertion of charged residues determine E degrees' changes that are invariably opposite to those induced by the concomitant enthalpic effects. Therefore, the resulting E degrees' changes are small or even opposite to those expected on simple electrostatic grounds. The mutational variation in the reduction entropy appears to be linked to the hydrogen bonding donor/acceptor character of the northern part of the protein, above the metal site, and to the electrostatic potential distribution around the copper site. Both properties influence the reduction-induced reorganization of the water molecules on the protein surface in the same region.

2001 - Electrostatic analysis and Brownian dynamics simulation of the association of plastocyanin and cytochrome F [Articolo su rivista]
DE RIENZO, Francesca; Rr, Gabdoulline; Menziani, Maria Cristina; DE BENEDETTI, Pier Giuseppe; Rc, Wade

The oxidation of cytochrome f by the soluble cupredoxin plastocyanin is a central reaction in the photosynthetic electron transfer chain of all oxygenic organisms. Here, two different computational approaches are used to gain new insights into the role of molecular recognition and protein-protein association processes in this redox reaction. First, a comparative analysis of the computed molecular electrostatic potentials of seven single and multiple point mutants of spinach plastocyanin (D42N, E43K, E43N, E43Q/D44N, E59K/E60Q, E59K/E60Q/E43N, Q88E) and the wt protein was carried out. The experimentally determined relative rates (k(2)) for the set of plastocyanin mutants are found to correlate well (r(2) = 0.90 - 0.97) with the computed measure of the similarity of the plastocyanin electrostatic potentials. Second, the effects on the plastocyanin/cytochrome f association rate of these mutations in the plastocyanin eastern site were evaluated by simulating the association of the wild type and mutant plastocyanins with cytochrome f by Brownian dynamics. Good agreement between the computed and experimental relative rates (k(2)) (r(2) = 0.89 - 0.92) was achieved for the plastocyanin mutants. The results obtained by applying both computational techniques provide support for the fundamental role of the acidic residues at the plastocyanin eastern site in the association with cytochrome f and in the overall electron-transfer process.

2001 - Enthalpic and entropic contributions to the mutational changes in the reduction potential of blue copper proteins [Abstract in Rivista]
Sola, Marco; Battistuzzi, Gianantonio; Borsari, Marco; G. W., Canters; E., de Waal; DE RIENZO, Francesca; Loschi, Lodovica; G., Warmerdam; Menziani, Maria Cristina

The changes in the reduction potential of Pseudomonas aeruginosa and Alcaligenes denitrificans azurins and spinachplastocyanin following point mutations and residue ionizations were factorized into the enthalpic and entropiccontributions through variable temperature direct electrochemistry experiments. The mutational variations of thereduction enthalpy and entropy were analyzed with a QSPR (Quantitative Structure-Property Relationships) approach,employing global and local theoretical descriptors defined and computed on the three dimensional protein structure.

2000 - Blue copper proteins: A comparative analysis of their molecular interaction properties [Articolo su rivista]
DE RIENZO, Francesca; Rr, Gabdoulline; Menziani, Maria Cristina; Rc, Wade

Blue copper proteins are type-I copper containing redox proteins whose role is to shuttle electrons from an electron donor to an electron acceptor in bacteria and plants. A large amount of experimental data is available on blue copper proteins; however, their functional characterization is hindered by the complexity of redox processes in biological systems. We describe here the application of a semiquantitative method based on a comparative analysis of molecular interaction fields to gain insights into the recognition properties of blue copper proteins. Molecular electrostatic and hydrophobic potentials were computed and compared for a set of 33 experimentally-determined structures of proteins from seven blue copper subfamilies, and the results were quantified by means of similarity indices. The analysis provides a classification of the blue copper proteins and shows that (1) comparison of the molecular electrostatic potentials provides useful information complementary to that highlighted by sequence analysis; (2) similarities in recognition properties can be detected for proteins belonging to different subfamilies, such as amicyanins and pseudoazurins, that may be isofunctional proteins; (3) dissimilarities in interaction properties, consistent with experimentally different binding specificities, may be observed between proteins belonging to the same subfamily, such as cyanobacterial and eukaryotic plastocyanins; (4) proteins with low sequence identity, such as azurins and pseudoazurins, can have sufficient similarity to bind to similar electron donors and accepters while having different binding specificity profiles.

2000 - Theoretical investigation of substrate specificity for cytochromes p450 IA2, p450 IID6 and p450 IIIA4 [Articolo su rivista]
DE RIENZO, Francesca; Fanelli, Francesca; Menziani, Maria Cristina; DE BENEDETTI, Pier Giuseppe

Three-dimensional models of the cytochromes P450 IA2, P450 IID6 and P450 IIIA4 were built by means of comparative modeling using the X-ray crystallographic structures of P450 CAM, P450 BM-3, P450 TERP and P450 ERYF as templates. The three cytochromes were analyzed both in their intrinsic structural features and in their interaction properties with fifty specific and non-specific substrates. Substrate/enzyme complexes were obtained by means of both automated rigid and flexible body docking. The comparative analysis of the three cytochromes and the selected substrates, in their free and bound forms, allowed for the building of semi-quantitative models of substrate specificity based on both molecular and intermolecular interaction descriptors. The results of this study provide new insights into the molecular determinants of substrate specificity for the three different eukaryotic P450 isozymes and constitute a useful tool for predicting the specificity of new compounds.