Submitted by Sara Ribeiro (sara.ribeiro@ucb.br) on 2017-11-09T10:26:59Z No. of bitstreams: 1 NicoleBerwangerCobachoDissertacao2017.pdf: 2681393 bytes, checksum: 06c28d06b89b63056ccfd84351dd8d36 (MD5) / Approved for entry into archive by Sara Ribeiro (sara.ribeiro@ucb.br) on 2017-11-09T11:17:54Z (GMT) No. of bitstreams: 1 NicoleBerwangerCobachoDissertacao2017.pdf: 2681393 bytes, checksum: 06c28d06b89b63056ccfd84351dd8d36 (MD5) / Made available in DSpace on 2017-11-09T11:17:55Z (GMT). No. of bitstreams: 1 NicoleBerwangerCobachoDissertacao2017.pdf: 2681393 bytes, checksum: 06c28d06b89b63056ccfd84351dd8d36 (MD5) Previous issue date: 2017-08-22 / UCB / The resistance of microorganisms to commonly used antibiotics has increased dramatically in recent years, suggesting that we will soon enter a post-antibiotic era where no therapy currently used will be effective in the fighting against infection. Thus, the search and study of new drugs and models of action of compounds that prevent or reduce the development of pathogens is essential. In this context, antimicrobial peptides (PAMs) appear as a new generation of compounds that demonstrate a great therapeutic potential. These molecules present themselves in low concentration in the organism of origin, making their isolation from natural sources an expensive and often impracticable process. In order to obtain a greater quantity of these peptides, recombinant expression via the heterologous system can be considered an efficient alternative in terms of time, cost and productivity. In 2009, Mandal et al. isolated three coconut water (Coco nucifera) peptides, called Cn-AMP1-2 and -3. Among them, Cn-AMP1 presented better activity against bacteria and fungi of medical importance. Subsequently, Lopez-Abarrategui et al. in 2012 isolated peptides with antimicrobial activity from the mollusk Cenchritis muricatus called Cm-p1 and Cm-p2. From these, a series of variant peptides were theoretically proposed in silico and evaluated against Candida albicans and the peptide named Cm-p5 demonstrated, among them, better antifungal activity. In this work, several constructs containing the peptides Cm-p5 and Cn-AMP1 were drawn in tandem and inserted into the pETSUMO (Life Technologies) vector. Genes were expressed in large quantities in strains derived from Escherichia coli BL21 (DE3) and after affinity column purification, they were evaluated, still fused with SUMO protein, against pathogenic microorganisms. However, none of the expressed peptides demonstrated bactericidal activity against the strains evaluated. / A resist??ncia dos microrganismos aos antibi??ticos que s??o comumente utilizados vem aumentando drasticamente nos ??ltimos anos, sugerindo que brevemente, entraremos em uma era p??s-antibi??ticos onde terapias utilizadas atualmente n??o ser??o mais eficientes no combate a infec????es. Desta forma, a procura e o estudo de novas drogas e modelos de a????o de compostos que impe??am ou reduzam o desenvolvimento de pat??genos ?? essencial. Neste contexto, os pept??deos antimicrobianos (PAMs) surgem como uma nova gera????o de compostos que demonstram um grande potencial terap??utico. Essas mol??culas apresentam-se em baixa concentra????o no organismo de origem, tornando o seu isolamento a partir de fontes naturais um processo dispendioso e, por muitas vezes, invi??vel. Para a obten????o de maior quantidade desses pept??deos, a express??o recombinante via sistema heter??logo pode ser considerada uma alternativa eficiente em rela????o ao tempo, custo e produtividade. Em 2009, Mandal e colaboradores isolaram tr??s pept??deos da agua de coco (Coco nucifera), denominados Cn-AMP1 -2 e -3. Dentre eles, o Cn-AMP1 apresentou melhor atividade contra bact??rias e fungos de import??ncia medica. Posteriormente, Lopez-Abarrategui e colaboradores em 2012 isolaram pept??deos com atividade antimicrobiana do molusco Centrichis muricatus denominados Cm-p1 e Cm-p2. A partir destes, uma serie de pept??deos variantes foram teoricamente propostos in silico e avaliados contra C. albicans sendo que o pept??deo nominado o Cm-p5 demonstrou, entre todos, melhor atividade antif??ngica. Neste trabalho, v??rias constru????es contendo os pept??deos Cm-p5 e Cn-AMP1 foram desenhadas em tandem e inseridas no vetor pETSUMO (Life Technologies). Os genes foram expressos em grandes quantidades em cepas derivadas de Escherichia coli BL21(DE3) e ap??s purifica????o em coluna de afinidade, os mesmos foram avaliados, ainda fusionados com a prote??na SUMO, contra microrganismos patog??nicos. No entanto, nenhum dos pept??deos expressos demonstrou atividade bactericida contra as cepas avaliadas.