Resposta de Chromobacterium Violaceum cultivada em alta concentra??o de ferro

• Main Author: Lima, Daniel Chaves de
• Other Authors: CPF:32398336468
• Format: Others
• Language: Portuguese
• Published: Universidade Federal do Rio Grande do Norte 2014
• Subjects: estresse; bottleneck; proteoma; biologia de sistemas; stress; proteome; systems biology; CNPQ::CIENCIAS BIOLOGICAS::BIOQUIMICA
• Online Access: http://repositorio.ufrn.br:8080/jspui/handle/123456789/12595

Made available in DSpace on 2014-12-17T14:03:39Z (GMT). No. of bitstreams: 1 DanielCL_DISSERT.pdf: 3446622 bytes, checksum: 86a8ec4f8ed7a90274cc61a7f0f71422 (MD5) Previous issue date: 2012-09-25 === Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior === The Chromobacterium violaceum is a β-proteobacterium Gram-negative widely found in tropical and subtropical regions, whose genome was sequenced in 2003 showing great metabolic versatility and biotechnological and pharmaceutical potential. Given the large number of ORFs related to iron metabolism described in the genome of C. violaceum, the importance of this metal for various biological processes and due to lack of data about the consequences of excess of iron in free-living organisms, it is important to study the response mechanism of this bacterium in a culture filled with iron. Previous work showed that C. violaceum is resistant to high concentrations of this metal, but has not yet been described the mechanism which is used to this survival. Thus, to elucidate the response of C. violaceum cultured in high concentrations of iron and expecting to obtain candidate genes for use in bioremediation processes, this study used a shotgun proteomics approach and systems biology to assess the response of C. violaceum grown in the presence and absence of 9 mM of iron. The analysis identified 531 proteins, being 71 exclusively expressed by the bacteria grown in the presence of the metal and 100 just in the control condition. The increase in expression of proteins related to the TCA cycle possibly represents a metabolic reprogramming of the bacteria caused by high concentration of iron in the medium. Moreover, we observed an increase in the activity assay of superoxide dismutase and catalase as well as in Total Antioxidant Activity assay, suggesting that the metal is inducing oxidative stress in C. violaceum that increases the levels of violacein and antioxidant enzymes to better adapt to the emerging conditions. Are also part of the adaptive response changes in expression of proteins related to transport, including iron, as well as an increased expression of proteins related to chemotaxis response, which would lead the bacteria to change the direction of its movement away from the metal. Systems Biology results, also suggest a metabolic reprogramming with mechanisms coordinated by bottleneck proteins involved in transcription (GreA), energy metabolism (Rpe and TpiA) and methylation (AhcY) === A Chromobacterium violaceum ? uma β-proteobact?ria Gram-negativa encontrada amplamente em regi?es tropicais e subtropicais, cujo genoma foi sequenciado em 2003 mostrando grande versatilidade metab?lica e potencial biotecnol?gico e farmac?utico. Dada ? grande quantidade de ORFs relacionadas com o metabolismo de ferro descritas no genoma da C. violaceum, a import?ncia deste metal para diversos processos biol?gicos e devido ? car?ncia de dados a respeito das conseq??ncias do excesso do ferro em organismos de vida livre, ? importante estudar o mecanismo de resposta desta bact?ria em meio repleto com ferro. Trabalhos anteriores mostram que a C. violaceum apresenta resist?ncia a grandes concentra??es desse metal, embora ainda n?o tenha sido descrito qual mecanismo ? utilizado para esta sobreviv?ncia. Assim, visando elucidar a resposta da C. violaceum cultivada em alta concentra??o de ferro e esperando-se obter genes candidatos para serem utilizados em processos de biorremedia??o, o presente trabalho utilizou a abordagem de prote?mica em shotgun e Biologia de Sistemas para avaliar a resposta de C. violaceum cultivada na presen?a e aus?ncia de 9 mM de ferro. A an?lise identificou 531 prote?nas, sendo 71 expressas exclusivamente pela bact?ria cultivada na presen?a do metal e 100 apenas na condi??o controle. O aumento na express?o de prote?nas relacionadas com o ciclo do TCA possivelmente representa uma reprograma??o metab?lica na bact?ria causada pela grande concentra??o de ferro no meio. Al?m disso, foi observado um aumento no ensaio de atividade das enzimas Super?xido dismutase e Catalase, bem como no ensaio de Atividade Antioxidante Total, sugerindo que o metal est? induzindo um quadro de estresse oxidativo em C. violaceum, que passou a aumentar os n?veis de violace?na e enzimas antioxidantes para melhor se adaptar ? condi??o emergente. Tamb?m faz parte da resposta adaptativa a altera??o na express?o de prote?nas relacionados a transporte, inclusive de ferro, e com a resposta quimiot?xica, o que levaria a bact?ria a mudar a orienta??o de sua locomo??o afastando-se do metal. Os dados de Biologia de Sistemas tamb?m sugerem uma reprograma??o metab?lica com mecanismos coordenados por gargalos proteicos envolvidos com transcri??o (GreA), metabolismo energ?tico (Rpe e TpiA) e metila??o (AhcY)