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Abstract: Short peptides can be designed in silico and synthesized through automated techniques, making them advantageous and versatile protein binders. A number of docking-based algorithms allow for a computational screening of peptides as binders. Here we developed ex-novo peptides targeting the maltose site of the Maltose Binding Protein, the prototypical system for the study of protein ligand recognition. We used a Monte Carlo based protocol, to computationally evolve a set of octapeptides starting from a polialanine sequence. We screened in silico the candidate peptides and characterized their binding abilities by surface plasmon resonance, fluorescence and electrospray ionization mass spectrometry assays. These experiments showed the designed binders to recognize their target with micromolar affinity. We finally discuss the obtained results in the light of further improvement in the ex-novo optimization of peptide based binders.
Found in: ključnih besedah
Keywords: peptides, docking, optimisation, computation, maltose binding protein, probe, ligand
Published: 12.10.2016; Views: 3262; Downloads: 132
Fulltext (4,27 MB)

Abstract: Tau is natively an unfolded protein that promotes the assembly and the stability of the axonal microtubules in the central nervous system. Increased formation of Tau protein aggregates has been causatively implicated in several neurodegenerative diseases called tauopathies. In the present study, we used the Drosophila melanogaster system to express the longest isoform of human Tau (2N4R) in the nervous system of adult flies, recreating the main features of the human pathology. Herein, this Tau-mediated neurodegeneration model was used as a platform to perform genetic screenings to identify putative modifiers of Tau toxicity. Our strategy exploited the modulation of genes considered as risk factors of Alzheimer’s disease (AD), Frontotemporal Dementias and other neurodegenerative diseases by RNA interference in vivo. This approach allowed us to identify a new gene which participates in the neuronal response against Tau induced neurotoxicity in Drosophila: D-Reep1, homologue of human REEP1 gene (h-Reep1). D-Reep1 knockout flies showed no apparent phenotypes in physiological growing and developmental conditions, however, they showed peculiar sensitivity to stress conditions. In addition, D-Reep1 knockout enhanced the neurodegeneration mediated by Tau expression in Drosophila eyes. On the contrary, the overexpression of UAS-D-Reep1 and UAS-h-Reep1 abolished the typical rough eye phenotype induced by the presence of Tau. The Co-expression of D-Reep1 in Tau backgrounds did not alter the phosphorylation pattern of this protein while, the presence of D-Reep1 seemed to prevent the formation of Tau aggregates in vivo. Thus, the data support the idea that D-Reep1 exerts a protective role on Tau induced toxicity which is independent of its phosphorylation status. In this work, I analysed the mechanisms behind the neuroprotective role of D-Reep1 and, in particular, I found that REEP1 is involved in the regulation of the unfolded protein response (UPR) through the PERK-ATF4 cascade within the ER. By the activation of this pathway, the neurotoxic aggregates of Tau are removed from Drosophila neuronal tissues rescuing the normal characteristics of the affected tissues. Evidences also suggest that the activation of autophagy was behind the removal of Tau aggregates, providing new molecular information about the physiological role of D Reep1 in the nervous system.
Found in: ključnih besedah
Summary of found: ...protein MT Microtubule/s MTBD Microtubule binding domain NFT Neurofibrillary tangle NP Neuritic plaques...
Keywords: AD Alzheimer Disease APP Amyloid precursor protein CNS Central Nervous System DM Drosophila melanogaster HSP Hereditary Spastic Paraplegia LN Lewy’s neurite MT Microtubule MAP Microtubule associated protein MT Microtubule/s MTBD Microtubule binding domain NFT Neurofibrillary tangle NP Neuritic plaques PHF Paired helical filament PS1 Presenilin 1 PS2 Presenilin 2 SPG Spastic Paraplegia ThS Thioflavin S
Published: 06.12.2019; Views: 2584; Downloads: 110
Fulltext (2,59 MB)

Found in: ključnih besedah
Summary of found: ... binding modes, drug delivery, drug release, iron oxide...
Keywords: binding modes, drug delivery, drug release, iron oxide nanoparticles, organic macrocycles, supramolecular interactions
Published: 02.09.2020; Views: 1661; Downloads: 0
Fulltext (3,39 MB)

Found in: ključnih besedah
Summary of found: ...crystal structures, DNA binding proteins, Methanocaldococcus jannaschii...
Keywords: crystal structures, DNA binding proteins, Methanocaldococcus jannaschii
Published: 16.03.2021; Views: 1264; Downloads: 8
Fulltext (0,00 KB)

Found in: ključnih besedah
Summary of found: ...DNA binding protein, Homo sapiens, synthetic constructs, Escherichia coli...
Keywords: DNA binding protein, Homo sapiens, synthetic constructs, Escherichia coli
Published: 17.03.2021; Views: 1432; Downloads: 7
Fulltext (0,00 KB)