Título:	Síntese química e caracterização das propriedades biológicas do peptídeo antimicrobiano Raniseptina-Bl 1 presente na secreção cutânea do anuro Boana lundii
Autor(es):	Silva, Brunna de Oliveira
Orientador(es):	Castro, Mariana de Souza
Assunto:	Anuros Boana lundii Peptídeos antimicrobianos Síntese química
Data de publicação:	29-Abr-2022
Data de defesa:	28-Dez-2021
Referência:	SILVA, Brunna de Oliveira. Síntese química e caracterização das propriedades biológicas do peptídeo antimicrobiano Raniseptina-Bl 1 presente na secreção cutânea do anuro Boana lundii. 2021. [77] f., il. Dissertação (Mestrado em Biologia Animal) — Universidade de Brasília, Brasília, 2021.
Abstract:	The indiscriminate use of antibiotics has contributed to the increase in the prevalence of pathogenic diseases promoted by multidrug-resistant microorganisms around the world, which represents a serious risk to human health. Given this situation, the search for new antimicrobial agents has mobilized the scientific community. Anurans have glands in their skin that produce and secrete a wide variety of bioactive compounds, including biologically active peptides. These molecules have an extensive range of bioactivities, such as antimicrobial, immunomodulatory, antiviral and antitumor activities. Antimicrobial peptides (AMPs) or host defense peptides (HDPs) have emerged as an alternative to face infections by multidrugresistant microorganims. The aim of the present work was to perform the chemical synthesis and characterization of the biological properties of the antimicrobial peptide Raniseptin-Bl 1 found in the cutaneous secretion of the anuran Boana lundii, an endemic species in Brazil. Raniseptin-Bl 1 was manually synthesized using the Fmoc/tBu method. The crude peptide was purified using RP-HPLC on a C18 column, resulting in the synthetic peptide with a high degree of purity, which was confirmed by MALDI-TOF mass spectrometry analysis, observing the presence of a single component with a protonated molecular mass equal to 2724.2 Da, corresponding to the native peptide isolated by Naywara Araújo, in 2018, from the cutaneous secretion of the anuran B. lundii. Raniseptin-Bl 1 represents the first peptide of the raniseptin family isolated from the species B. lundii exhibiting high identity and similarity indices with Raniseptins 3 and 13, previously identified in the skin of the anuran B. raniceps. Circular dichroism analysis, in the presence of SDS micelles, showed that Raniseptin-Bl 1 assumed a predominant secondary structure conformation in α-helix, with a helicity of 82.7%. The synthetic raniseptin-Bl 1 was tested against 4 bacteria and 2 species of pathogenic fungi. The peptide was effective in inhibiting the growth of Gram-positive bacteria with MIC values of 4 µM for Staphylococcus aureus and 2 µM for S. epidermidis. In the case of the Gram-negative bacteria tested, a higher activity of Raniseptin-Bl 1 was observed, with values of 0.5 µM for Escherichia coli and 1 µM for Klebsiella pneumoniae. The peptide was also effective in inhibiting the growth of pathogenic yeasts Candida albicans with a MIC value of 8 µM and Cryptococcus neoformans with a MIC equal to 64 µM. Its ability to lyse human erythrocytes was also evaluated, showing a moderate hemolysis rate, less than 40%, at the highest concentration used (128 M). In the region of the MICs values determined for the tested bacteria (0.5–4 M), the percentage of hemolysis was below 10%, indicating a low cytolytic activity. The morphological changes suffered by the Gram-negative bacteria E. coli treated with raniseptin-Bl 1 were evaluated by scanning electron microscopy (SEM). Bacteria treated at different concentrations (2, 4 and 8 µM) showed roughness, irregularities, and deformities on the cell surface. We also analyzed the effects of the peptide Raniseptin Bl 1 on innate immunity by evaluating its influence on the modulation of the phagocytic capacity and the production of NETs by human neutrophils. The incubation of neutrophils with Raniseptin-Bl 1, despite apparently influencing the result, did not have statistical validation in the case of phagocytic activity and NET production. There is an apparent interaction between Raniseptin-Bl 1 and fMLP in the production of NETs, however there was no statistical confirmation by 2-way ANOVA of the central point. Analyzing the straight line parameters, there is a significant difference in the linear coefficient between quiescent neutrophils and the combination of Raniseptin-Bl 1 and fMLP, which suggests a possible combined action, perhaps synergistic between them, in a way that together they are probably able to induce the formation of NETs. Raniseptin-Bl 1 exhibited promising antimicrobial properties, with MIC values ranging from 0.5 to 8 µM for the microorganisms tested, in addition to a low hemolytic activity (less than 10% hemolysis in the range detected for MICs). This set of results signals the potential of Raniseptin-Bl 1 to serve as a model for the development of new antimicrobial agents in order to overcome the current crisis associated with microbial resistance to commercially available antibiotics.
Unidade Acadêmica:	Instituto de Ciências Biológicas (IB)
Informações adicionais:	Dissertação (mestrado) — Universidade de Brasília, Instituto de Ciências Biológicas, Programa de Pós-Graduação em Biologia Animal, 2021.
Programa de pós-graduação:	Programa de Pós-Graduação em Biologia Animal
Licença:	A concessão da licença deste item refere-se ao termo de autorização impresso assinado pelo autor com as seguintes condições: Na qualidade de titular dos direitos de autor da publicação, autorizo a Universidade de Brasília e o IBICT a disponibilizar por meio dos sites www.bce.unb.br, www.ibict.br, http://hercules.vtls.com/cgi-bin/ndltd/chameleon?lng=pt&skin=ndltd sem ressarcimento dos direitos autorais, de acordo com a Lei nº 9610/98, o texto integral da obra disponibilizada, conforme permissões assinaladas, para fins de leitura, impressão e/ou download, a título de divulgação da produção científica brasileira, a partir desta data.
Agência financiadora:	Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES).
Aparece nas coleções:	Teses, dissertações e produtos pós-doutorado

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