One single method to produce native and Tat-fused recombinant human α-synuclein in Escherichia coli

Caldinelli, Laura; Albani, Diego; Pollegioni, Loredano

doi:10.1186/1472-6750-13-32

Cited by 19 publications

(15 citation statements)

References 38 publications

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“…The effects of antisynuclein (anti‐syn) and anti‐Aβ antibodies (4G8) on memory impairment induced by intracerebroventricular application of alpha‐synuclein oligomers (α‐synOs) and determined by the novel object recognition test. The antibodies (0.25 μg) were applied 15 minutes before the oligomers (1 μM of recombinant synuclein) dissolved in PBS (Vehicle). After a habituation period, mice were exposed for 10 minutes to 2 identical objects (familiar).…”

Section: In Vivo Application Of Aβ and α‐Syn Oligomers: Differences Amentioning

confidence: 99%

Oligomeropathies and pathogenesis of Alzheimer and Parkinson's diseases

et al. 2016

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The term oligomeropathies defines the neurodegenerative disorders associated with protein misfolding, where small soluble aggregates (oligomers 4-200 KDa) are the cause of neuronal dysfunction and are responsible for spreading the pathology. The ability of these soluble β-sheet conformers to induce neuronal damage has been investigated in direct challenge with the monomeric and fibrillary structures, showing that only the oligomeric species affected the neurons. β amyloid oligomers were initially purified from Alzheimer brains and obtained using synthetic peptides. Together with the neuronal death, synaptic dysfunction, loss of spines, and LTP impairment were seen with the direct application of β amyloid oligomers. Similar results have been described with proteins associated with other neurodegenerative disorders. The biological activities of oligomeric forms of α synuclein have been described in Parkinson's disease and Lewy body dementia. Detrimental effects have been associated with the oligomeric forms of prion, tau, and huntingtin, the key proteins in prion diseases, frontotemporal dementia, and Huntington's disease, respectively. The molecular mechanisms of the oligomer-related toxic effects can be summarized under three headings: nonspecific perturbance of cellular and intracellular membranes, specific interaction with various cellular entities, and amyloid pore channel formation. To characterize and distinguish oligomer actions better, we compared the ability of β amyloid and α synuclein oligomers to induce cognitive impairment when applied directly into the brain in the same acute mouse model. We also investigated the role of inflammatory components. © 2016 International Parkinson and Movement Disorder Society.

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Section: In Vivo Application Of Aβ and α‐Syn Oligomers: Differences Amentioning

confidence: 99%

Oligomeropathies and pathogenesis of Alzheimer and Parkinson's diseases

et al. 2016

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“…Recombinant α‐Syn and TAT‐α‐Syn were expressed and purified from periplasmic space content as reported in Caldinelli et al [19]. The pET5a‐α‐Syn plasmid was a gift of Prof. Bartels (Boston, Massachusetts, United states of America).…”

Section: Methodsmentioning

confidence: 99%

Insight into conformational modification of alpha‐synuclein in the presence of neuronal whole cells and of their isolated membranes

et al. 2015

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a b s t r a c tA change in the conformational plasticity of a-Synuclein (a-Syn) is hypothesised to be a key step in the pathogenic mechanism of Parkinson's disease (PD). Here, we report the study of extracellular aSyn interaction with whole cells and membranes isolated from the neuronal SH-SY5Y cells, exploiting NMR and CD spectroscopies. In addition, the crosslinking agent DSG was used to freeze the conformational and oligomeric state of a-Syn in the presence of cells. These data, in a quasi-physiological environment, confirm the protein monomeric state with a propensity to adopt a transient alpha helical following interaction with biological membranes.

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“…For protein expression, the recombinant pNIC28-Bsa4 vector containing αS gene was transformed into E. coli C41 (DE3); starter cultures were prepared by inoculating a single c olony of the bacteria to LB medium containing kanamycin (50 μg.mL -1 ). After reaching to an of OD 600nm 0.4, IPTG (0.1 mM) was added, and the culture was incubated for another 5 hours at 37°C (22). The induced cells were harvested by centrifugation at 5000 ×g for 15 min.…”

Section: Expression and Purification Of αSmentioning

confidence: 99%

TiO2 Nanoparticles as Potential Promoting Agents of Fibrillation of α-Synuclein, a Parkinson’s Disease-Related Protein

Mohammadi

Nikkhah

2017

Iran. J. Biotechnol.

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In recent years, nanomaterials have been widely used in large quantities which make people be more frequently exposed to the chemically synthesized nanoparticles (NPs). When NPs are introduced into an organism, they may interact with a variety of cellular components with yet largely unknown pathological consequences. It was found that NPs enhance the rate of protein fibrillation in the brain by decreasing the lag time for nucleation. Protein fibrillation is implicated in the pathogenesis of the several neurodegenerative diseases such as Parkinson's disease (PD). α-Synuclein (αS) is natively an unfolded protein which is involved in the pathogenesis of PD. In the present study, we have analyzed the effects of three different NPs on αS fibrillation. αS protein expression and purifi cation was done and fibrils formation was induced in the absence or presence of the three types of NPs (i. e., TiO2, SiO2, and SnO2). The enhancement of the fluorescence emission of Thiofl avin T (ThT) and transmission electron microscopy (TEM) were used to monitor the appearance and growth of the fibrils. The adsorption of αS monomers on the surface of NPs was investigated by tyrosine fluorescence emission measurements. We found that TiO2-NPs enhances αS fibril formation even at a concentration of 5 μg.mL, while the two other NPs show no significant effect on the kinetics of the fibrillation. Intrinsic tyrosine emission measurement has confirmed that the TiO2-NPs interact with αS fibrillation products. It is suggested that TiO2- NPs may enhance the nucleation of αS protein that leads to protein fibril formation. The fibrillization process of αS protein is profoundly affected by the presence of TiO2-NPs. This finding unveils the neurotoxicity potential of the TiO2-NPs, which may be considered as a probable risk for PD.

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One single method to produce native and Tat-fused recombinant human α-synuclein in Escherichia coli

Cited by 19 publications

References 38 publications

Oligomeropathies and pathogenesis of Alzheimer and Parkinson's diseases

Oligomeropathies and pathogenesis of Alzheimer and Parkinson's diseases

Insight into conformational modification of alpha‐synuclein in the presence of neuronal whole cells and of their isolated membranes

TiO2 Nanoparticles as Potential Promoting Agents of Fibrillation of α-Synuclein, a Parkinson’s Disease-Related Protein

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