Claudia Andreini

University of Florence | UNIFI · Dipartimento di Chimica "Ugo Schiff"

Metalloproteins are ubiquitous in all kingdoms of life. Their role and function are tightly related to the local structure of the metal-binding site. In this regard, the MetalPDB database is an invaluable tool since it stores the 3D structure of metal-binding sites and of their corresponding apo forms. In this work, we exploited MetalPDB to compute...

All living organisms require metal ions for their energy production and metabolic and biosynthetic processes. Within cells, the metal ions involved in the formation of adducts interact with metabolites and macromolecules (proteins and nucleic acids). The proteins that require binding to one or more metal ions in order to be able to carry out their...

Iron-sulfur (Fe/S) clusters are protein cofactors that play a crucial role in essential cellular functions. Their ability to rapidly exchange electrons with several redox active acceptors makes them an efficient system for fulfilling diverse cellular needs. They include the formation of a relay for long-range electron transfer in enzymes, the biosy...

The Protein Data Bank in Europe – Knowledge Base (PDBe-KB, https://pdbe-kb.org) is an open collaboration between world-leading specialist data resources contributing functional and biophysical annotations derived from or relevant to the Protein Data Bank (PDB). The goal of PDBe-KB is to place macromolecular structure data in their biological contex...

Previous work has shown that the Tat protein of Human Immunodeficiency Virus (HIV)-1 is released by acutely infected cells in a biologically active form and enters dendritic cells upon the binding of its arginine-glycine-aspartic acid (RGD) domain to the α5β1, αvβ3, and αvβ5 integrins. The up-regulation/activation of these integrins occurs in endot...

We developed a novel force field in the context of AMBER parameterization for glutamate and aspartate zinc(II)-binding residues. The interaction between the zinc ion and the coordinating atoms is represented by a spherical nonbonded parameterization. The polarization effect due to the zinc ion has been taken into account by redefining the atomic ch...

Iron-sulfur clusters are ubiquitous protein cofactors composed of iron and inorganic sulfur. These cofactors are among the most ancient ones and may have contributed to the birth of life on Earth. Therefore, they are found even today in many enzymes central to metabolic processes like nitrogen fixation, respiration, and DNA processing and repair. D...

We developed and validated a novel force field in the context of the AMBER parameterization for the simulation of zinc(II)-binding proteins. The proposed force field assumes non-bonded spherical interactions between the central zinc(II) and the coordinating residues. A crucial innovative aspect of our approach is to account for the polarization eff...

Calprotectin (CP) inhibits bacterial viability through extracellular chelation of transition metals. However, how CP influences general metabolism remains largely unexplored. We show here that CP restricts bioavailable Zn and Fe to the pathogen Acinetobacter baumannii, inducing an extensive multi-metal perturbation of cellular physiology. Proteomic...

Zinc is an essential cofactor for many proteins. A key mechanism of zinc homeostasis during deficiency is “zinc sparing” in which specific zinc-binding proteins are repressed to reduce the cellular requirement. In this report, we evaluated zinc sparing across the zinc proteome of Saccharomyces cerevisiae. The yeast zinc proteome of 582 known or pot...

Organisms from all kingdoms of life use iron-proteins in a multitude of functional processes. We applied a bioinformatics approach to investigate the human portfolio of iron-proteins. We separated iron-proteins based on the chemical nature of their metal-containing cofactors: individual iron ions, heme cofactors and iron–sulfur clusters. We found t...

Superfamilies without splitting events. This table lists superfamilies that contain enzymes all associated to a single EC number.

Iron, zinc, and copper play fundamental roles in eucaryotes and procaryotes, and their bioavailability regulates host-pathogen interactions. For intracellular pathogens, the source of metals is the cytoplasm of the host, which in turn manipulates intracellular metal traffic following pathogen recognition. It is established that iron is withheld fro...

3-D reconstruction of AX2 cells infected with GFP-producing L. pneumophila 48 hpi. 3-D projections of a single AX2 cell infected with GFP-tagged L. pneumophila were generated and animated with the confocal microscope LSM800 software (Zen Software, Carl Zeiss, Inc., Oberkochen, Germany) in.avi format. The movie shows bacteria contained in high numbe...

Plaque formation by AX2 cells previously grown in media depleted or overloaded with iron, zinc, or copper. Ax2 cells were grown in M1 ± Fe for 24 h or in M2 ± Zn or M3 ± Cu for 3 weeks. Cells were then washed, resuspended in Soerensen buffer, and serial dilutions were plated on a lawn of E. coli B2, S. typhimurium, or K. aerogenes, as described in...

About half of known enzymatic reactions involve metals. Enzymes belonging to the same superfamily often evolve to catalyze different reactions on the same structural scaffold. The work presented here investigates how functional differentiation, within superfamilies that contain metalloenzymes, relates to structural changes at the catalytic metal si...

MetalPDB (http://metalweb.cerm.unifi.it/) is a database providing information on metal-binding sites detected in the three-dimensional (3D) structures of biological macromolecules. MetalPDB represents such sites as 3D templates, called Minimal Functional Sites (MFSs), which describe the local environment around the metal(s) independently of the lar...

Tetrahymena thermophila (T. thermophila) is a ciliated protozoon that can detect freshwater pollution by heavy metals (“whole-cell biosensor”). This work employed a systematic bioinformatics approach to predict and analyze the metalloproteome of T. thermophila for the essential Zn, Cu and the non-essential Cd. 3784 metal-binding proteins were ident...

About 2 billion years ago, the atmosphere of the Earth experienced a great change due to the buildup of dioxygen produced by photosynthetic organisms. This transition caused a reduction of iron bioavailability and at the same time exposed living organisms to the threat of oxidative stress. Iron-sulfur (Fe-S) clusters require iron ions for their bio...

Occurrence in aerobes of the Fe-S families specific to HC anaerobes. For each Fe-S family specific of HC anaerobes (i.e. conserved in at least 30% the HC anaerobes and in less than 30% aerobes) the graph shows the percentage of aerobic organisms which (i) do not have a corresponding family member (grey); (ii) have members without a Fe-S-binding dom...

Families (COGs) that correspond to the predicted Fe-S proteins and analysis of the frequently occurring Fe-S families in HC anaerobes and aerobes. (XLSX)

Occurrence in HC anaerobes of the Fe-S families specific to aerobes. For each Fe-S family specific of aerobes (i.e. conserved in at least 30% aerobes and in less than 30% HC anaerobes) is reported the percentage of HC anaerobic organisms which (i) do not have a corresponding family member (grey); (ii) have members without a Fe-S-binding domain (gre...

(A) Percentage of aerobic organisms that encode a given Fe-S family (y axis) as a function of the percentage of obligate aerobic organisms that encode the same family (x axis). (B) Percentage of aerotolerant HC organisms that encode a given Fe-S family (y axis) as a function of the percentage of obligate anaerobic organisms that encode the same fam...

Fe-S proteins predicted in obligate aerobes, aerobes, facultative anaerobes, aerotolerant anaerobes and obligate anaerobes. (XLSX)

Families (COGs) that correspond to the 70 predicted Fe-S proteins in humans and their mapping to families of aerobic and HC anaerobic organisms. (XLSX)

Average number of co-orthologs found in Fe-S families of the shared core. Blue columns are for aerobes; red columns are for and anaerobes. (TIF)

Number of Fe-S proteins and Fe-S families per each organism analyzed (obligate aerobes, aerobes, facultative anaerobes, aerotolerant anaerobes and obligate anaerobes are grouped in separate tabs). (XLSX)

Motivation: The prediction of the iron–sulfur proteome is highly desirable for biomedical and biological research but a freely available tool to predict iron–sulfur proteins has not been developed yet. Results: We developed a web server to predict iron–sulfur proteins from protein sequence(s). This tool, called MetalPredator, is able to process co...

Metal ions play a functional role in numerous biochemical processes and cellular pathways. Indeed, about 40% of all enzymes of known 3D structure require a metal ion to be able to perform catalysis. The interactions of the metals with the macromolecular framework determine their chemical properties and reactivity. The relevant interactions involve...

Organisms from all kingdoms of life use Iron-Sulfur Proteins (ISPs) in a multitude of functional processes. We applied a bioinformatics approach to investigate the human portfolio of ISPs. 61% of human ISPs bind Fe4S4 clusters whereas 39% bind Fe2S2 clusters. However, this relative ratio varies significantly depending on the specific cellular compa...

In this work, we developed a methodology to perform a systematic classification based on three-dimensional structural similarity of the metal sites contained in metalloproteins. Our definition of metal site extended beyond the metal ion and its aminoacidic ligands by including all the chemical species (aminoacids, nucleotides, exogenous ligands) pr...

Metalloproteins account for a substantial fraction of all proteins. They incorporate metal atoms, which are required for their structure and/or function. Here we describe a new computational protocol to systematically compare and classify metal-binding sites on the basis of their structural similarity. These sites are extracted from the MetalPDB da...

We have developed a database search tool to identify metal sites having structural similarity to a query metal site structure within the MetalPDB database of minimal functional sites (MFSs) contained in metal-binding biological macromolecules. MFSs describe the local environment around the metal(s) independently of the larger context of the macromo...

We developed a new software tool, MetalS(2), for the structural alignment of Minimal Functional Sites (MFSs) in metal-binding biological macromolecules. MFSs are 3D templates that describe the local environment around the metal(s) independently of the larger context of the macromolecular structure. Such local environment has a determinant role in t...

We present here MetalPDB (freely accessible at http://metalweb.cerm.unifi.it), a novel resource aimed at conveying the information available on the three-dimensional (3D) structures of metal-binding biological macromolecules in a consistent and effective manner. This is achieved through the systematic and automated representation of metal-binding s...

Use of Env in HIV vaccine development has been disappointing. Here we show that, in the presence of a biologically active Tat subunit vaccine, a trimeric Env protein prevents in monkeys virus spread from the portal of entry to regional lymph nodes. This appears to be due to specific interactions between Tat and Env spikes that form a novel virus en...

Entry of wt Tat in MDDCs in the presence of different Env molecules and block by anti-integrins antibodies. Clade B trimeric wt Env (twt Env), trimeric ΔV2 Env (tΔV2 Env), monomeric wt Env (mwt Env), or monomeric ΔV2 Env (mΔV2 Env) molecules or buffer were incubated with Tat and added to MDDCs pre-treated with anti-integrin mAbs or a control isotyp...

Structural model of the ΔV1-2 Env/Tat binary complex. Color code: Env: dark blue; Env V3-loop: light blue; Tat: yellow; Tat cysteine-rich region: orange; and Tat RGD segment: red. (A) panels: surface representation; (B) panels: cartoon representation. See experimental procedures for details. (TIF)

Blockade of Tat/Env complex entry into MDDCs by anti-Tat antibodies. Trimeric ΔV2 Env was incubated with PBS, a pool of sera from six HIV uninfected healthy donors, the same pool of sera plus the anti-Tat 2A4.1 mAb, Tat, or Tat plus 2A4.1 mAb, and added to cells. Cells were then stained for intracellular Env and analyzed by flow cytometry. The perc...

Blockade of Tat/Env complex binding in PBL by anti-CD4 antibodies. PBLs were pre-incubated with buffer or an anti-CD4 mAb and then incubated with twt Env or with twt Env which had been pre-incubated with the indicated amounts of Tat. PBLs were then stained with an anti-gp120 mAb and analyzed by flow cytometry. The percentage of Env-binding cells is...

Wt Tat, but not cys22 Tat, transactivates HIV-1 LTR in TZM-bl cells. RLU in HIV-1 LTR expressing TZM-bl cells cultured in the presence of buffer, or 0.1, 1, 10 µM wt Tat or cys22 Tat. (TIF)

Block of trimeric ΔV2 Env or Tat/ΔV2 Env entry in MDDCs by anti-DC-SIGN and anti-integrins antibodies. (A) MDDCs from two different donors were pre-incubated with different concentration of an anti-DC-SIGN mAb (20 or 50 µg/mL) or with a control isotype mAb and then 70 nM (donor 1) or 35 nM (donor 2) trimeric ΔV2 Env (SF162) were added for 10 min pr...

Env and Tat interacting residues according to modeling docking analyses. Upper panel: Env interacting residues in the five lowest energy solutions. Residues involved in interactions are indicated by boxes. Different box colors correspond to different solutions. Secondary structure elements are colored as follows: the ΔV1-2 gp120 inner domain of Env...

Vaccine protocol design and schedule of immunization of cynomolgus monkeys. (DOCX)

Structures used as templates to model the structures of Tat and Env. (DOC)

Parameters used to perform MD simulations on the V3 loop of the Env protein. (DOC)

Parameters used to perform docking calculations. (DOC)

Parameters used to perform docking calculations. (DOC)

Structural Model of the ΔV1-2 Env/Tat/Integrin αvβ3 Ternary Complex. Color code: ΔV1-2 Env: violet; Tat: yellow; integrin αvβ3: cyan. (TIF)

CD4+ T cell counts, plasma viral load and proviral DNA load in blood, inguinal lymph nodes and rectal mucosal tissues at 4 week after intrarectal challenge with 70 MID50 of SHIVSF162P4cy.

Unlabelled: Metals are essential for the structure and function of many proteins and nucleic acids. The geometrical arrangement of the atoms that coordinate a metal in a biological macromolecule is an important determinant of the specificity and role of that metal. At present, however, this information can be retrieved only from the literature, wh...

Thanks to the contributions of scientists like Bert Vallee, zinc enzymology is an area of research with a rich history and a strong basis of biochemical and biophysical knowledge. In recent years, the dramatic development of the genomic and post-genomic research has provided this as well as all other fields of life sciences with a massive body of n...

MACiE (which stands for Mechanism, Annotation and Classification in Enzymes) is a database of enzyme reaction mechanisms, and can be accessed from http://www.ebi.ac.uk/thornton-srv/databases/MACiE/. This article presents the release of Version 3 of MACiE, which not only extends the dataset to 335 entries, covering 182 of the EC sub-subclasses with...

Results of the clustering of representative Zn-sites using different distance threshold values for defining spatially proximal residues in building the MFS templates. (PDF)

Summary of the relevant information on Zn-superfamilies. (PDF)

Lists of the Zn-sites belonging to each Zn-superfamily. (PDF)

Schematic picture of the structures of the representative Zn-sites included in each Zn-cluster. (PDF)

List of the non-physiological Zn-sites found in PDB structures and removed from the dataset. (PDF)

Zinc is indispensable to all forms of life as it is an essential component of many different proteins involved in a wide range of biological processes. Not differently from other metals, zinc in proteins can play different roles that depend on the features of the metal-binding site. In this work, we describe zinc sites in proteins with known struct...

A biochemical pathway can be viewed as a series of chemical reactions occurring within a cell, each of which is carried out by one or more biological macromolecules (protein, RNA, or complexes thereof). Computational methods can be applied to assess whether one organism is able to perform a biochemical process of interest by checking whether its ge...

Genome-wide studies are providing researchers with a potentially complete list of the molecular components present in living systems. It is now evident that several metal ions are essential to life and that metalloproteins, that is, proteins that require a metal ion to perform their physiological function, are widespread in all organisms. However,...

Metal-MACiE is a new publicly available web-based database, held in MySQL, which aims to organize the available information on the properties and the roles of metals in the context of the catalytic mechanisms of metalloenzymes. Metal-MACiE, which currently covers 75% of metal-dependent enzyme commission (EC) sub-sub-classes and is continuously grow...

In metalloproteins, the protein environment modulates metal properties to achieve the required goal, which can be protein stabilization or function. The analysis of metal sites at the atomic level of detail provided by protein structures can thus be of benefit in functional and evolutionary studies of proteins. In this work, we propose a structural...

We analysed the roles and distribution of metal ions in enzymatic catalysis using available public databases and our new resource Metal-MACiE (http://www.ebi.ac.uk/thornton-srv/databases/Metal_MACiE/home.html). In Metal-MACiE, a database of metal-based reaction mechanisms, 116 entries covering 21% of the metal-dependent enzymes and 70% of the types...

In high-throughput genome-level protein investigation efforts, such as Structural Genomics, the systematic experimental characterization of metal-binding properties (i.e., the investigation of the metalloproteome) is not always pursued and remains far from trivial. In the present work, we have applied a bioinformatic approach to investigate the occ...

A novel ArsR-SmtB family transcriptional repressor, KmtR, has been characterized from mycobacteria. Mutants of Mycobacterium tuberculosis lacking kmtR show elevated expression of Rv2025c encoding a deduced CDF-family metal exporter. KmtR-dependent repression of the cdf and kmtR operator-promoters was alleviated by nickel and cobalt in minimal mediu...

Metalloproteins are proteins capable of binding one or more metal ions, which are often required for their biological function or for regulation of their activities or for structural purposes. In high-throughput genome-level protein investigation efforts, such as Structural Genomics, the systematic experimental characterization of metal-binding pro...

SPCH-SVM vs previous methods. Comparisons at a chain level between predictions of the SPCH-SVM and results published in Andreini et al., J. Proteome Research, 5(1):196–201, 2006.

SVM evaluation data. List of protein chains employed in the experiments, splits of the 5-fold cross validation procedure, model parameters obtained in the tuning phase.

[RefSeq:NP_060357.1] alignment. Metal binding pattern alignment for [RefSeq:NP_060357.1].

Metalloproteins are proteins capable of binding one or more metal ions, which may be required for their biological function, for regulation of their activities or for structural purposes. Metal-binding properties remain difficult to predict as well as to investigate experimentally at the whole-proteome level. Consequently, the current knowledge abo...

Zinc is one of the metal ions essential for life, as it is required for the proper functioning of a large number of proteins. Despite its importance, the annotation of zinc-binding proteins in gene banks or protein domain databases still has significant room for improvement. In the present work, we compiled a list of known zinc-binding protein doma...

SPINE (Structural Proteomics In Europe) was established in 2002 as an integrated research project to develop new methods and technologies for high-throughput structural biology. Development areas were broken down into workpackages and this article gives an overview of ongoing activity in the bioinformatics workpackage. Developments cover target sel...

http://www.cerm.unifi.it/ Abstract. We describe and empirically evaluate machine learning methods for the prediction of zinc binding sites from protein sequences. We start by observing that a data set consisting of single residues as ex- amples is affected by autocorrelation and we propose an ad-hoc remedy in which sequentially close pairs of candi...

Many bacterial genomes encode multiple metal-sensing ArsR-SmtB transcriptional repressors. There is interest in understanding and predicting their metal specificities. Here we analyse two arsR-smtB genes, ydeT and yozA (now aseR and czrA) from Bacillus subtilis. Purified AseR and CzrA formed complexes in gel-retardation and fluorescence-anisotropy...

The present bioinformatic research attempts to answer the question: how many and which proteins encoded in the human genome require zinc for their function? It is found that around 10% of all human proteins are zinc-binding proteins. Zinc has a crucial role in stabilizing the structure of transcription factors, next to its widespread use in all enz...

The "A Disintegrin And Metalloproteinase" (ADAM) protein family and the "A Disintegrin-like And Metalloproteinase with ThromboSpondin motifs" (ADAMTS) protein family are two related families of human proteins. The similarities and differences between these two families have been investigated using phylogenetic trees and homology modeling. The phylo...

With the advent of genome sequencing, a huge database of protein primary sequences has been accumulating. In parallel, a number of tools to investigate and expand upon this information, e.g. reconstructing and building relationships between protein families and superfamilies, have been developed. Metalloproteins are proteins capable of binding one...

The entire family of human matrix metalloproteinases (MMPs) was investigated using phylogenetic trees and homology modeling. The phylogenetic analysis indicates that individual domains of each MMP have evolved in a correlated manner. Despite their high sequence similarity, the phylogenetic tree of the catalytic domains already allows functional (e....