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1.
Nanoscale ; 10(19): 9174-9185, 2018 May 17.
Artigo em Inglês | MEDLINE | ID: mdl-29725687

RESUMO

The protein α-synuclein (αSN) aggregates to form fibrils in neuronal cells of Parkinson's patients. Here we report on the effect of neutral (zwitterionic) nanoliposomes (NLPs), supplemented with cholesterol (NLP-Chol) and decorated with PEG (NLP-Chol-PEG), on αSN aggregation and neurotoxicity. Both NLPs retard αSN fibrillization in a concentration-independent fashion. They do so largely by increasing lag time (formation of fibrillization nuclei) rather than elongation (extension of existing nuclei). Interactions between neutral NLPs and αSN may locate to the N-terminus of the protein. This interaction can even perturb the interaction of αSN with negatively charged NLPs which induces an α-helical structure in αSN. This interaction was found to occur throughout the fibrillization process. Both NLP-Chol and NLP-Chol-PEG were shown to be biocompatible in vitro, and to reduce αSN neurotoxicity and reactive oxygen species (ROS) levels with no influence on intracellular calcium in neuronal cells, emphasizing a prospective role for NLPs in reducing αSN pathogenicity in vivo as well as utility as a vehicle for drug delivery.


Assuntos
Lipossomos/química , Nanopartículas/química , Neurônios/efeitos dos fármacos , Doença de Parkinson/terapia , alfa-Sinucleína/química , 1,2-Dipalmitoilfosfatidilcolina/química , Animais , Cálcio/metabolismo , Colesterol/química , Humanos , Células PC12 , Polietilenoglicóis/química , Ratos , Espécies Reativas de Oxigênio/metabolismo
2.
Protein Expr Purif ; 129: 75-83, 2017 01.
Artigo em Inglês | MEDLINE | ID: mdl-27664437

RESUMO

Vessel dilator is a 3.9-KDa potent anticancer peptide and a valuable candidate in the treatment of conditions such as congestive heart failure and acute renal failure amongst others. Here we report the recombinant production of vessel dilator in Escherichia coli. Three different synthetic ORF's dubbed VDI, VDII and VDIII, each encoding a trimmer of the vessel dilator peptide attached to a His tag sequence at their C- terminal, were synthesized and placed in pET21c expression vectors. The highest yield, following expression in E. coli BL21 (DE3), was recorded with VDII that carried the shortest fusion partner. Subsequent to the initial capture of the fusion protein by a Ni affinity column, the vessel dilator monomers were cleaved by trypsin treatment, and further purified to at least 90% homogeneity by anion exchange chromatography. De-novo sequencing and in vivo anticancer activity tests were used to verify the peptide sequence and its biological activity, respectively. The final yield was estimated to be approximately 15 mg of the purified vessel dilator per gram wet weight of the bacterial cells.


Assuntos
Antineoplásicos , Fator Natriurético Atrial , Neoplasias Colorretais/tratamento farmacológico , Escherichia coli/metabolismo , Antineoplásicos/isolamento & purificação , Antineoplásicos/metabolismo , Antineoplásicos/farmacologia , Fator Natriurético Atrial/biossíntese , Fator Natriurético Atrial/genética , Fator Natriurético Atrial/isolamento & purificação , Fator Natriurético Atrial/farmacologia , Linhagem Celular Tumoral , Cromatografia de Afinidade , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Escherichia coli/genética , Histidina/biossíntese , Histidina/isolamento & purificação , Humanos , Fragmentos de Peptídeos/biossíntese , Fragmentos de Peptídeos/genética , Fragmentos de Peptídeos/isolamento & purificação , Fragmentos de Peptídeos/farmacologia , Proteínas Recombinantes de Fusão/biossíntese , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/isolamento & purificação , Proteínas Recombinantes de Fusão/farmacologia
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