Navegando por Palavras-chave "Catepsina A"

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    Identificação de potenciais antígenos do carrapato Rhipicephalus (Boophilus) microplus para o desenvolvimento de vacinas
    (Universidade Federal de São Paulo (UNIFESP), 2020-07-30) Silva, Fernando Allan Abreu [UNIFESP]; Tanaka, Aparecida Sadae [UNIFESP]; Universidade Federal de São Paulo
    The tick Rhipicephalus (Boophilus) microplus is responsible for considerable economic losses to the livestock sector. The main control method of infestation of this parasite is by acaricides and can result in the selection of resistant ticks, environment and products contamination, between other factors. The vaccine is a more sustainable alternative, but the products available on the market are not efficient in the Brazilian territory. Although the tick genome has not yet been elucidated, transcriptomes and proteomes can indicant potential targets for selection of antigens. Two proteins sequences were selected for this work, a sphingomyelinase D – like and a cathepsin A – like from a tick transcriptome. The DNA sequences that codified these two proteins were cloned into the pET14b expression vector and confirmed by DNA sequencing. The analysis of the translated amino acids of cathepsin A – like showed the absence of amino acids residues important for the activity of this enzyme when compared to others cathepsins A sequences. However, it was not possible to confirm experimentally this data since the recombinant protein was not express in bacterial system. In contrast, sphingomyelinase D – like was expressed in bacteria in large quantities, but in the form of inclusion body. The sphingomyelinase D – like was partially purified in the presence of urea. The analysis of sphingomyelinase D – like primary structure showed a replacement of a histidine for an asparagine in an important position for the enzymatic activity. Therefore, the perspective of this work is to obtain the purified sphingomyelinase D – like and carry out its biochemical and functional characterization.
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    Papel de miRNAs na regulação da expressão do complexo multienzimático lisossômico
    (Universidade Federal de São Paulo (UNIFESP), 2017-04-28) Queiroz, Matheus Trovão de [UNIFESP]; D'Almeida, Vania [UNIFESP]; Pereira, Vanessa Gonçalves [UNIFESP]; http://lattes.cnpq.br/4126672233843478; http://lattes.cnpq.br/7220411418339421; http://lattes.cnpq.br/2833936812464737; Universidade Federal de São Paulo (UNIFESP)
    Mucopolysaccharidosis type I (MPS I) is a genetic disease in which there is a deficiency in the expression of the lysosomal enzyme α-L-iduronidase (IDUA gene), which culminates in a generalized lysosomal disturbance. Thus, in addition to the storage of direct substrates of α-L-iduronidase, it is observed the involvement of secondary pathways such as the increase of simple gangliosides levels in the central nervous system, implicated in the neurodegenerative process observed in this disease. Recent studies have shown that Neu1 gene (neuraminidase 1 enzyme), a member of the lysosomal multienzyme complex (CML) involved in ganglioside metabolism, is significantly reduced in the cerebellum and hippocampus of the murine MPS I model. Recently, alterations in the levels of three miRNAs from miR-17 family were reported, which only miR-20b-5p presents as predicted direct targets Neu1 and Ctsa genes, the latter encoding cathepsin A enzyme, another important member of CML. Aiming the functional characterization of miR-20b-5p, the involvement of this miRNA with the regulation of neuraminidase 1 expression and the observed effects on gene and protein expression of CML members in a context of miR-20b- 5p overexpression in cell culture were evaluated. It was observed miR-20b- 5p is not involved with the regulation of Neu1 expression by its 3' untranslated region. In addition, obtained data suggest that this miRNA is involved in fine tuning RNAm levels of the CLM members, but it has no implications for the level of protein expression. Thus, these results suggest the existence of a complex genetic regulation circuit of CML and that miR-20b-5p may represent an important piece in this context.