| Texto completo | |
| Autor(es): |
Santos, Joao H. P. M.
;
Feitosa, Valker A.
;
Meneguetti, Giovanna P.
;
Carretero, Gustavo
;
Coutinho, Joao A. P.
;
Ventura, Sonia P. M.
;
Rangel-Yagui, Carlota O.
Número total de Autores: 7
|
| Tipo de documento: | Artigo Científico |
| Fonte: | BIOSENSORS-BASEL; v. 12, n. 2, p. 13-pg., 2022-02-01. |
| Resumo | |
Cytochrome c (Cyt-c), a small mitochondrial electron transport heme protein, has been employed in bioelectrochemical and therapeutic applications. However, its potential as both a biosensor and anticancer drug is significantly impaired due to poor long-term and thermal stability. To overcome these drawbacks, we developed a site-specific PEGylation protocol for Cyt-c. The PEG derivative used was a 5 kDa mPEG-NHS, and a site-directed PEGylation at the lysine amino-acids was performed. The effects of the pH of the reaction media, molar ratio (Cyt-c:mPEG-NHS) and reaction time were evaluated. The best conditions were defined as pH 7, 1:25 Cyt-c:mPEG-NHS and 15 min reaction time, resulting in PEGylation yield of 45% for Cyt-c-PEG-4 and 34% for Cyt-c-PEG-8 (PEGylated cytochrome c with 4 and 8 PEG molecules, respectively). Circular dichroism spectra demonstrated that PEGylation did not cause significant changes to the secondary and tertiary structures of the Cyt-c. The long-term stability of native and PEGylated Cyt-c forms was also investigated in terms of peroxidative activity. The results demonstrated that both Cyt-c-PEG-4 and Cyt-c-PEG-8 were more stable, presenting higher half-life than unPEGylated protein. In particular, Cyt-c-PEG-8 presented great potential for biomedical applications, since it retained 30-40% more residual activity than Cyt-c over 60-days of storage, at both studied temperatures of 4 degrees C and 25 degrees C. (AU) | |
| Processo FAPESP: | 16/22065-5 - Pegilação N-terminal de proteínas e purificação por sistemas aquosos bifásicos |
| Beneficiário: | Carlota de Oliveira Rangel Yagui |
| Modalidade de apoio: | Auxílio à Pesquisa - Regular |
| Processo FAPESP: | 18/25994-2 - Desenvolvimento de novas plataformas de PEGuilação de proteínas com potencial terapêutico com recurso à microfluídica |
| Beneficiário: | João Henrique Picado Madalena Santos |
| Modalidade de apoio: | Bolsas no Brasil - Pós-Doutorado |