18 results on '"Giorgi, Alessandra"'
Search Results
2. β-Hexachlorocyclohexane triggers neuroinflammatory activity, epigenetic histone post-translational modifications and cognitive dysfunction
- Author
-
Grieco, Maddalena, Giorgi, Alessandra, Giacovazzo, Giacomo, Maggiore, Anna, Ficchì, Serena, d'Erme, Maria, Mosca, Luciana, Mignogna, Giuseppina, Maras, Bruno, and Coccurello, Roberto
- Published
- 2024
- Full Text
- View/download PDF
3. Rab11A Depletion in Microglia-Derived Extracellular Vesicle Proteome upon Beta-Amyloid Treatment
- Author
-
Mignogna, Giuseppina, Fabrizi, Cinzia, Correani, Virginia, Giorgi, Alessandra, and Maras, Bruno
- Published
- 2023
- Full Text
- View/download PDF
4. Ferritin‐based disruptor nanoparticles: A novel strategy to enhance LDL cholesterol clearance via multivalent inhibition of PCSK9–LDL receptor interaction.
- Author
-
Incocciati, Alessio, Cappelletti, Chiara, Masciarelli, Silvia, Liccardo, Francesca, Piacentini, Roberta, Giorgi, Alessandra, Bertuccini, Lucia, De Berardis, Barbara, Fazi, Francesco, Boffi, Alberto, Bonamore, Alessandra, and Macone, Alberto
- Abstract
Hypercholesterolemia, characterized by elevated low‐density lipoprotein (LDL) cholesterol levels, is a significant risk factor for cardiovascular disease. Proprotein convertase subtilisin/kexin type 9 (PCSK9) plays a crucial role in cholesterol metabolism by regulating LDL receptor degradation, making it a therapeutic target for mitigating hypercholesterolemia‐associated risks. In this context, we aimed to engineer human H ferritin as a scaffold to present 24 copies of a PCSK9‐targeting domain. The rationale behind this protein nanoparticle design was to disrupt the PCSK9–LDL receptor interaction, thereby attenuating the PCSK9‐mediated impairment of LDL cholesterol clearance. The N‐terminal sequence of human H ferritin was engineered to incorporate a 13‐amino acid linear peptide (Pep2‐8), which was previously identified as the smallest PCSK9 inhibitor. Exploiting the quaternary structure of ferritin, engineered nanoparticles were designed to display 24 copies of the targeting peptide on their surface, enabling a multivalent binding effect. Extensive biochemical characterization confirmed precise control over nanoparticle size and morphology, alongside robust PCSK9‐binding affinity (KD in the high picomolar range). Subsequent efficacy assessments employing the HepG2 liver cell line demonstrated the ability of engineered ferritin's ability to disrupt PCSK9–LDL receptor interaction, thereby promoting LDL receptor recycling on cell surfaces and consequently enhancing LDL uptake. Our findings highlight the potential of ferritin‐based platforms as versatile tools for targeting PCSK9 in the management of hypercholesterolemia. This study not only contributes to the advancement of ferritin‐based therapeutics but also offers valuable insights into novel strategies for treating cardiovascular diseases. [ABSTRACT FROM AUTHOR]
- Published
- 2024
- Full Text
- View/download PDF
5. A cross-cultural analysis of the gestural pattern of surprise and surprise-disapproval questions.
- Author
-
Giorgi, Alessandra and Petrocchi, Erika
- Subjects
CROSS-cultural studies ,SURPRISE ,LANGUAGE & languages ,GESTURE ,VIETNAMESE language - Abstract
In this article, we address the issue concerning the gestural patterns in expressing surprise and disapproval across various languages and cultures. The results obtained so far point to an interesting, and in a sense rather surprising, uniformity. We consider two types of special questions: counter-expectational questions expressing surprise and surprise-disapproval questions, i.e., sentences expressing surprise with a negative orientation, and adopt an experimental design involving sentence repetition and spontaneous production. We focus on the realization of these sentences in Vietnamese, Korean and Japanese, which we compare with the results previously obtained for Italian and replicated for Neapolitan, Spanish and German. Our research is based on the Minimalist theoretical framework developed by Chomsky and scholars in the tradition of generative grammar. [ABSTRACT FROM AUTHOR]
- Published
- 2024
- Full Text
- View/download PDF
6. Ferritin nanocage-enabled detection of pathological tau in living human retinal cells.
- Author
-
Barolo, Lorenzo, Gigante, Ylenia, Mautone, Lorenza, Ghirga, Silvia, Soloperto, Alessandro, Giorgi, Alessandra, Ghirga, Francesca, Pitea, Martina, Incocciati, Alessio, Mura, Francesco, Ruocco, Giancarlo, Boffi, Alberto, Baiocco, Paola, and Di Angelantonio, Silvia
- Subjects
FERRITIN ,INDUCED pluripotent stem cells ,MEDICAL sciences ,TAU proteins ,ALZHEIMER'S disease ,TAUOPATHIES ,NANOMEDICINE ,POLYMERSOMES - Abstract
Tauopathies, including Alzheimer's disease and Frontotemporal Dementia, are debilitating neurodegenerative disorders marked by cognitive decline. Despite extensive research, achieving effective treatments and significant symptom management remains challenging. Accurate diagnosis is crucial for developing effective therapeutic strategies, with hyperphosphorylated protein units and tau oligomers serving as reliable biomarkers for these conditions. This study introduces a novel approach using nanotechnology to enhance the diagnostic process for tauopathies. We developed humanized ferritin nanocages, a novel nanoscale delivery system, designed to encapsulate and transport a tau-specific fluorophore, BT1, into human retinal cells for detecting neurofibrillary tangles in retinal tissue, a key marker of tauopathies. The delivery of BT1 into living cells was successfully achieved through these nanocages, demonstrating efficient encapsulation and delivery into retinal cells derived from human induced pluripotent stem cells. Our experiments confirmed the colocalization of BT1 with pathological forms of tau in living retinal cells, highlighting the method's potential in identifying tauopathies. Using ferritin nanocages for BT1 delivery represents a significant contribution to nanobiotechnology, particularly in neurodegenerative disease diagnostics. This method offers a promising tool for the early detection of tau tangles in retinal tissue, with significant implications for improving the diagnosis and management of tauopathies. This study exemplifies the integration of nanotechnology with biomedical science, expanding the frontiers of nanomedicine and diagnostic techniques. [ABSTRACT FROM AUTHOR]
- Published
- 2024
- Full Text
- View/download PDF
7. Combined Delivery of miR-15/16 through Humanized Ferritin Nanocages for the Treatment of Chronic Lymphocytic Leukemia.
- Author
-
Liberati, Francesca Romana, Di Russo, Sara, Barolo, Lorenzo, Peruzzi, Giovanna, Farina, Maria Vittoria, Spizzichino, Sharon, Di Fonzo, Federica, Quaglio, Deborah, Pisano, Luca, Botta, Bruno, Giorgi, Alessandra, Boffi, Alberto, Cutruzzolà, Francesca, Paone, Alessio, and Baiocco, Paola
- Subjects
CHRONIC lymphocytic leukemia ,FERRITIN ,B cell lymphoma ,B cells - Abstract
Chronic lymphocytic leukemia (CLL) is a widespread type of leukemia that predominantly targets B lymphocytes, undermining the balance between cell proliferation and apoptosis. In healthy B cells, miR-15/16, a tandem of microRNAs, functions as a tumor suppressor, curbing the expression of the antiapoptotic B cell lymphoma 2 protein (Bcl-2). Conversely, in CLL patients, a recurring deletion on chromosome 13q14, home to the miR15-a and miR16-1 genes, results in Bcl-2 overexpression, thereby fostering the onset of the pathology. In the present research, a novel approach utilizing humanized ferritin-based nanoparticles was employed to successfully deliver miR15-a and miR-16-1 into MEG01 cells, a model characterized by the classic CLL deletion and overexpression of the human ferritin receptor (TfR1). The loaded miR15-a and miR16-1, housed within modified HumAfFt, were efficiently internalized via the MEG01 cells and properly directed into the cytoplasm. Impressively, the concurrent application of miR15-a and miR16-1 demonstrated a robust capacity to induce apoptosis through the reduction in Bcl-2 expression levels. This technology, employing RNA-loaded ferritin nanoparticles, hints at promising directions in the battle against CLL, bridging the substantial gap left by traditional transfection agents and indicating a pathway that may offer hope for more effective treatments. [ABSTRACT FROM AUTHOR]
- Published
- 2024
- Full Text
- View/download PDF
8. Identification of the pyridoxal 5′‐phosphate allosteric site in human pyridox(am)ine 5′‐phosphate oxidase.
- Author
-
Barile, Anna, Graziani, Claudio, Antonelli, Lorenzo, Parroni, Alessia, Fiorillo, Annarita, di Salvo, Martino Luigi, Ilari, Andrea, Giorgi, Alessandra, Rosignoli, Serena, Paiardini, Alessandro, Contestabile, Roberto, and Tramonti, Angela
- Abstract
Adequate levels of pyridoxal 5′‐phosphate (PLP), the catalytically active form of vitamin B6, and its proper distribution in the body are essential for human health. The PLP recycling pathway plays a crucial role in these processes and its defects cause severe neurological diseases. The enzyme pyridox(am)ine 5′‐phosphate oxidase (PNPO), whose catalytic action yields PLP, is one of the key players in this pathway. Mutations in the gene encoding PNPO are responsible for a severe form of neonatal epilepsy. Recently, PNPO has also been described as a potential target for chemotherapeutic agents. Our laboratory has highlighted the crucial role of PNPO in the regulation of PLP levels in the cell, which occurs via a feedback inhibition mechanism of the enzyme, exerted by binding of PLP at an allosteric site. Through docking analyses and site‐directed mutagenesis experiments, here we identified the allosteric PLP binding site of human PNPO. This site is located in the same protein region as the allosteric site we previously identified in the Escherichia coli enzyme homologue. However, the identity and arrangement of the amino acid residues involved in PLP binding are completely different and resemble those of the active site of PLP‐dependent enzymes. The identification of the PLP allosteric site of human PNPO paves the way for the rational design of enzyme inhibitors as potential anti‐cancer compounds. [ABSTRACT FROM AUTHOR]
- Published
- 2024
- Full Text
- View/download PDF
9. Pragmatics in the Minimalist framework: Evidence from the study of emotional language.
- Author
-
Giorgi, Alessandra
- Subjects
PRAGMATICS ,LARGE-scale brain networks ,POINT processes ,LANGUAGE & languages ,GRAMMAR - Abstract
This article explores the relationship between pragmatics and the other components of grammar. Specifically, it aims to determine whether pragmatics is a distinct module of grammar coming into play at some point in the derivation process to connect the sentence with the context. The conclusion is that, based on the phenomena considered in this work, pragmatics rather than being a separate module, is distributed in the various components. It is shown in fact that the context immediately intervenes at the representative level to yield the correct syntax to be fed to the sensorimotor system on one side, and to the conceptual one on the other. The empirical focus of the article is on a specific type of questions in Italian, namely surprise and surprise-disapproval questions, because they are most sensitive to pragmatic factors. The syntactic, prosodic, and gestural components of these constructions will be examined, highlighting their most important characteristics. [ABSTRACT FROM AUTHOR]
- Published
- 2023
- Full Text
- View/download PDF
10. Proteomic characterization of extracellular vesicles released by third stage larvae of the zoonotic parasite Anisakis pegreffii (Nematoda: Anisakidae).
- Author
-
Palomba, Marialetizia, Rughetti, Aurelia, Mignogna, Giuseppina, Castrignanò, Tiziana, Rahimi, Hassan, Masuelli, Laura, Napoletano, Chiara, Pinna, Valentina, Giorgi, Alessandra, Santoro, Mario, Schininà, Maria Eugenia, Maras, Bruno, and Mattiucci, Simonetta
- Subjects
EXTRACELLULAR vesicles ,PARASITES ,PROTEOMICS ,ANISAKIS ,LIFE cycles (Biology) ,LARVAE ,NEMATODES ,CETACEA - Abstract
Introduction: Anisakis pegreffii is a sibling species within the A. simplex (s.l.) complex requiring marine homeothermic (mainly cetaceans) and heterothermic (crustaceans, fish, and cephalopods) organisms to complete its life cycle. It is also a zoonotic species, able to accidentally infect humans (anisakiasis). To investigate the molecular signals involved in this host-parasite interaction and pathogenesis, the proteomic composition of the extracellular vesicles (EVs) released by the third-stage larvae (L3) of A. pegreffii, was characterized. Methods: Genetically identified L3 of A. pegreffii were maintained for 24 h at 37° C and EVs were isolated by serial centrifugation and ultracentrifugation of culture media. Proteomic analysis was performed by Shotgun Analysis. Results and discussion: EVs showed spherical shaped structure (size 65-295 nm). Proteomic results were blasted against the A. pegreffii specific transcriptomic database, and 153 unique proteins were identified. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analysis predicted several proteins belonging to distinct metabolic pathways. The similarity search employing selected parasitic nematodes database revealed that proteins associated with A. pegreffii EVs might be involved in parasite survival and adaptation, as well as in pathogenic processes. Further, a possible link between the A. pegreffii EVs proteins versus those of human and cetaceans' hosts, were predicted by using HPIDB database. The results, herein described, expand knowledge concerning the proteins possibly implied in the host-parasite interactions between this parasite and its natural and accidental hosts. [ABSTRACT FROM AUTHOR]
- Published
- 2023
- Full Text
- View/download PDF
11. Structural Characterization of Murine Phosphodiesterase 5 Isoforms and Involvement of Cysteine Residues in Supramolecular Assembly.
- Author
-
Giorgi, Mauro, Miele, Adriana Erica, Cardarelli, Silvia, Giorgi, Alessandra, Massimi, Mara, Biagioni, Stefano, and Saliola, Michele
- Subjects
POLYACRYLAMIDE gel electrophoresis ,SMALL-angle X-ray scattering ,KLUYVEROMYCES marxianus ,CYSTEINE ,TRANSMISSION electron microscopy ,QUATERNARY structure ,POLYACRYLAMIDE - Abstract
Phosphodiesterases (PDEs) are a superfamily of evolutionarily conserved cyclic nucleotide (cAMP/cGMP)-hydrolyzing enzymes, components of transduction pathways regulating crucial aspects of cell life. Within this family, the cGMP-dependent PDE5 is the major hydrolyzing enzyme in many mammalian tissues, where it regulates a number of cellular and tissular processes. Using Kluyveromyces lactis as a model organism, the murine PDE5A1, A2 and A3 isoforms were successfully expressed and studied, evidencing, for the first time, a distinct role of each isoform in the control, modulation and maintenance of the cellular redox metabolism. Moreover, we demonstrated that the short N-terminal peptide is responsible for the tetrameric assembly of MmPDE5A1 and for the mitochondrial localization of MmPDE5A2. We also analyzed MmPDE5A1, A2 and A3 using small-angle X-ray scattering (SAXS), transmission electron microscopy (TEM), structural mass spectrometry (MS) and polyacrylamide gel electrophoresis in their native conditions (native-PAGE) and in the presence of redox agents. These analyses pointed towards the role of a few specific cysteines in the isoforms' oligomeric assembly and the loss of enzymatic activity when modified. [ABSTRACT FROM AUTHOR]
- Published
- 2023
- Full Text
- View/download PDF
12. Multicomponent Synthesis of Diaminopurine and Guanine PNA's Analogues Active against Influenza A Virus from Prebiotic Compounds.
- Author
-
Bizzarri, Bruno Mattia, Fanelli, Angelica, Ciprini, Stefania, Giorgi, Alessandra, De Angelis, Marta, Fioravanti, Raoul, Nencioni, Lucia, and Saladino, Raffaele
- Published
- 2022
- Full Text
- View/download PDF
13. Modulation of Virulence-Associated Traits in Aspergillus fumigatus by BET Inhibitor JQ1.
- Author
-
Orekhova, Anastasia, De Angelis, Marta, Cacciotti, Andrea, Reverberi, Massimo, Rotili, Dante, Giorgi, Alessandra, Protto, Virginia, Bonincontro, Graziana, Fiorentino, Francesco, Zgoda, Victor, Mai, Antonello, Palamara, Anna Teresa, and Simonetti, Giovanna
- Subjects
ASPERGILLUS fumigatus ,MELANINS ,GREATER wax moth ,ANTIFUNGAL agents ,MELANOGENESIS ,SUPEROXIDE dismutase ,LACCASE - Abstract
Aspergillus fumigatus is a disease-causing, opportunistic fungus that can establish infection due to its capacity to respond to a wide range of environmental conditions. Secreted proteins and metabolites, which play a critical role in fungal–host interactions and pathogenesis, are modulated by epigenetic players, such as bromodomain and extraterminal domain (BET) proteins. In this study, we evaluated the in vitro and in vivo capability of the BET inhibitor JQ1 to modulate the extracellular proteins and virulence of A. fumigatus. The abundance of 25 of the 76 extracellular proteins identified through LC-MS/MS proteomic analysis changed following JQ1 treatment. Among them, a ribonuclease, a chitinase, and a superoxide dismutase were dramatically downregulated. Moreover, the proteomic analysis of A. fumigatus intracellular proteins indicated that Abr2, an intracellular laccase involved in the last step of melanin synthesis, was absent in the JQ1-treated group. To investigate at which level this downregulation occurred and considering the ability of JQ1 to modulate gene expression we checked the level of ABR2, Chitinase, and Superoxide dismutase mRNA expression by qRT-PCR. Finally, the capacity of JQ1 to reduce the virulence of A. fumigatus has been proved using Galleria mellonella larvae, which are an in vivo model to evaluate fungal virulence. Overall, the promising activity exhibited by JQ1 suggests that A. fumigatus is sensitive to BET inhibition and BET proteins may be a viable target for developing antifungal agents. [ABSTRACT FROM AUTHOR]
- Published
- 2022
- Full Text
- View/download PDF
14. Characterization of the Escherichia coli pyridoxal 5′‐phosphate homeostasis protein (YggS): Role of lysine residues in PLP binding and protein stability.
- Author
-
Tramonti, Angela, Ghatge, Mohini S., Babor, Jill T., Musayev, Faik N., di Salvo, Martino Luigi, Barile, Anna, Colotti, Gianni, Giorgi, Alessandra, Paredes, Steven D., Donkor, Akua K., Al Mughram, Mohammed H., de Crécy‐Lagard, Valérie, Safo, Martin K., and Contestabile, Roberto
- Abstract
The pyridoxal 5′‐phosphate (PLP) homeostasis protein (PLPHP) is a ubiquitous member of the COG0325 family with apparently no catalytic activity. Although the actual cellular role of this protein is unknown, it has been observed that mutations of the PLPHP encoding gene affect the activity of PLP‐dependent enzymes, B6 vitamers and amino acid levels. Here we report a detailed characterization of the Escherichia coli ortholog of PLPHP (YggS) with respect to its PLP binding and transfer properties, stability, and structure. YggS binds PLP very tightly and is able to slowly transfer it to a model PLP‐dependent enzyme, serine hydroxymethyltransferase. PLP binding to YggS elicits a conformational/flexibility change in the protein structure that is detectable in solution but not in crystals. We serendipitously discovered that the K36A variant of YggS, affecting the lysine residue that binds PLP at the active site, is able to bind PLP covalently. This observation led us to recognize that a number of lysine residues, located at the entrance of the active site, can replace Lys36 in its PLP binding role. These lysines form a cluster of charged residues that affect protein stability and conformation, playing an important role in PLP binding and possibly in YggS function. PDB Code(s): 7U9C, 7U9H, 7UAT, 7UAU, 7UAX, 7UBP, 7UBA, 7UB8 and 7UBQ; [ABSTRACT FROM AUTHOR]
- Published
- 2022
- Full Text
- View/download PDF
15. Failed Directives and Fatal Commissives : Speech Act Analysis of Oscar Wilde’s Salome
- Author
-
Kurzon, Dennis, Capone, Alessandro, Series Editor, Goldberg, Sanford, Advisory Editor, Graci, Roberto, Advisory Editor, Macagno, Fabrizio, Advisory Editor, Sharvit, Yael, Advisory Editor, Allan, Keith, Advisory Editor, Cummings, Louise, Advisory Editor, Davis, Wayne A., Advisory Editor, Douven, Igor, Advisory Editor, Kecskes, Istvan, Advisory Editor, Pennisi, Antonino, Advisory Editor, Santuli, Francesca, Advisory Editor, Burton-Roberts, Noel, Editorial Board Member, Butler, Brian, Editorial Board Member, Carapezza, Marco, Editorial Board Member, Cimatti, Felice, Editorial Board Member, Corazza, Eros, Editorial Board Member, Devitt, Michael, Editorial Board Member, van Eemeren, Frans, Editorial Board Member, Feit, Neil, Editorial Board Member, Giorgi, Alessandra, Editorial Board Member, Horn, Larry, Editorial Board Member, von Heusinger, Klaus, Editorial Board Member, Jaszczolt, Kasia, Editorial Board Member, Jeshion, Robin Beth, Editorial Board Member, Korta, Kepa, Editorial Board Member, Lepore, Ernest, Editorial Board Member, Levinson, Stephen C., Editorial Board Member, Piazza, Francesca, Editorial Board Member, Richard, Mark, Editorial Board Member, Salmon, Nathan, Editorial Board Member, Schiffer, Stephen R., Editorial Board Member, Seymour, Michel, Editorial Board Member, Simons, Mandy, Editorial Board Member, Williamson, Timothy, Editorial Board Member, Wierbizcka, Anna, Editorial Board Member, Traugott, Elizabeth C., Editorial Board Member, and Perconti, Pietro, editor
- Published
- 2024
- Full Text
- View/download PDF
16. Ferritin-based disruptor nanoparticles: A novel strategy to enhance LDL cholesterol clearance via multivalent inhibition of PCSK9-LDL receptor interaction.
- Author
-
Incocciati A, Cappelletti C, Masciarelli S, Liccardo F, Piacentini R, Giorgi A, Bertuccini L, De Berardis B, Fazi F, Boffi A, Bonamore A, and Macone A
- Subjects
- Humans, PCSK9 Inhibitors pharmacology, PCSK9 Inhibitors chemistry, Ferritins chemistry, Ferritins metabolism, Protein Binding, Proprotein Convertase 9 metabolism, Proprotein Convertase 9 chemistry, Proprotein Convertase 9 genetics, Receptors, LDL metabolism, Receptors, LDL chemistry, Nanoparticles chemistry, Cholesterol, LDL metabolism
- Abstract
Hypercholesterolemia, characterized by elevated low-density lipoprotein (LDL) cholesterol levels, is a significant risk factor for cardiovascular disease. Proprotein convertase subtilisin/kexin type 9 (PCSK9) plays a crucial role in cholesterol metabolism by regulating LDL receptor degradation, making it a therapeutic target for mitigating hypercholesterolemia-associated risks. In this context, we aimed to engineer human H ferritin as a scaffold to present 24 copies of a PCSK9-targeting domain. The rationale behind this protein nanoparticle design was to disrupt the PCSK9-LDL receptor interaction, thereby attenuating the PCSK9-mediated impairment of LDL cholesterol clearance. The N-terminal sequence of human H ferritin was engineered to incorporate a 13-amino acid linear peptide (Pep2-8), which was previously identified as the smallest PCSK9 inhibitor. Exploiting the quaternary structure of ferritin, engineered nanoparticles were designed to display 24 copies of the targeting peptide on their surface, enabling a multivalent binding effect. Extensive biochemical characterization confirmed precise control over nanoparticle size and morphology, alongside robust PCSK9-binding affinity (K
D in the high picomolar range). Subsequent efficacy assessments employing the HepG2 liver cell line demonstrated the ability of engineered ferritin's ability to disrupt PCSK9-LDL receptor interaction, thereby promoting LDL receptor recycling on cell surfaces and consequently enhancing LDL uptake. Our findings highlight the potential of ferritin-based platforms as versatile tools for targeting PCSK9 in the management of hypercholesterolemia. This study not only contributes to the advancement of ferritin-based therapeutics but also offers valuable insights into novel strategies for treating cardiovascular diseases., (© 2024 The Author(s). Protein Science published by Wiley Periodicals LLC on behalf of The Protein Society.)- Published
- 2024
- Full Text
- View/download PDF
17. Identification of the pyridoxal 5'-phosphate allosteric site in human pyridox(am)ine 5'-phosphate oxidase.
- Author
-
Barile A, Graziani C, Antonelli L, Parroni A, Fiorillo A, di Salvo ML, Ilari A, Giorgi A, Rosignoli S, Paiardini A, Contestabile R, and Tramonti A
- Subjects
- Humans, Allosteric Site, Phosphates, Pyridoxal Phosphate metabolism, Oxidoreductases metabolism, Pyridoxaminephosphate Oxidase genetics, Pyridoxaminephosphate Oxidase metabolism
- Abstract
Adequate levels of pyridoxal 5'-phosphate (PLP), the catalytically active form of vitamin B
6 , and its proper distribution in the body are essential for human health. The PLP recycling pathway plays a crucial role in these processes and its defects cause severe neurological diseases. The enzyme pyridox(am)ine 5'-phosphate oxidase (PNPO), whose catalytic action yields PLP, is one of the key players in this pathway. Mutations in the gene encoding PNPO are responsible for a severe form of neonatal epilepsy. Recently, PNPO has also been described as a potential target for chemotherapeutic agents. Our laboratory has highlighted the crucial role of PNPO in the regulation of PLP levels in the cell, which occurs via a feedback inhibition mechanism of the enzyme, exerted by binding of PLP at an allosteric site. Through docking analyses and site-directed mutagenesis experiments, here we identified the allosteric PLP binding site of human PNPO. This site is located in the same protein region as the allosteric site we previously identified in the Escherichia coli enzyme homologue. However, the identity and arrangement of the amino acid residues involved in PLP binding are completely different and resemble those of the active site of PLP-dependent enzymes. The identification of the PLP allosteric site of human PNPO paves the way for the rational design of enzyme inhibitors as potential anti-cancer compounds., (© 2024 The Authors. Protein Science published by Wiley Periodicals LLC on behalf of The Protein Society.)- Published
- 2024
- Full Text
- View/download PDF
18. Characterization of the Escherichia coli pyridoxal 5'-phosphate homeostasis protein (YggS): Role of lysine residues in PLP binding and protein stability.
- Author
-
Tramonti A, Ghatge MS, Babor JT, Musayev FN, di Salvo ML, Barile A, Colotti G, Giorgi A, Paredes SD, Donkor AK, Al Mughram MH, de Crécy-Lagard V, Safo MK, and Contestabile R
- Subjects
- Lysine metabolism, Pyridoxal Phosphate, Proteins chemistry, Protein Stability, Homeostasis, Phosphates metabolism, Carrier Proteins genetics, Escherichia coli metabolism, Escherichia coli Proteins genetics, Escherichia coli Proteins metabolism
- Abstract
The pyridoxal 5'-phosphate (PLP) homeostasis protein (PLPHP) is a ubiquitous member of the COG0325 family with apparently no catalytic activity. Although the actual cellular role of this protein is unknown, it has been observed that mutations of the PLPHP encoding gene affect the activity of PLP-dependent enzymes, B
6 vitamers and amino acid levels. Here we report a detailed characterization of the Escherichia coli ortholog of PLPHP (YggS) with respect to its PLP binding and transfer properties, stability, and structure. YggS binds PLP very tightly and is able to slowly transfer it to a model PLP-dependent enzyme, serine hydroxymethyltransferase. PLP binding to YggS elicits a conformational/flexibility change in the protein structure that is detectable in solution but not in crystals. We serendipitously discovered that the K36A variant of YggS, affecting the lysine residue that binds PLP at the active site, is able to bind PLP covalently. This observation led us to recognize that a number of lysine residues, located at the entrance of the active site, can replace Lys36 in its PLP binding role. These lysines form a cluster of charged residues that affect protein stability and conformation, playing an important role in PLP binding and possibly in YggS function., (© 2022 The Authors. Protein Science published by Wiley Periodicals LLC on behalf of The Protein Society.)- Published
- 2022
- Full Text
- View/download PDF
Catalog
Discovery Service for Jio Institute Digital Library
For full access to our library's resources, please sign in.