Title:	Proteômica top-down da bactéria produtora de aminoácidos Corynebacterium glutamicum
Authors:	Melo, Reynaldo Magalhães
Orientador(es)::	Vale, Luis Henrique Ferreira do
Assunto::	Corynebacterium glutamicum Aminoácidos - produção Microorganismos Bactérias
Issue Date:	22-Aug-2019
Data de defesa::	27-Feb-2019
Citation:	MELO, Reynaldo Magalhães. Proteômica top-down da bactéria produtora de aminoácidos Corynebacterium glutamicum. 2019. 116 f., il. Dissertação (Mestrado em Biologia Microbiana)—Universidade de Brasília, Brasília, 2019.
Abstract:	Corynebacterium glutamicum is a workhorse bacterium used to produce many biomolecules, among them there are strains capable to produce isobutanol, succinate, panthothenate, biopolimers and other compounds. Amino acids are the most explored group of molecules for industrial production in C. glutamicum. These are the third most produced biomolecules through fermentation worldwide. The C. glutamicum genome sequencing was published in 2003, and it increased our capacity to genetically modify it. Among the “omics”, proteomics presents great potential to characterize metabolic pathways, once it is able to identify PTMs that are required for metabolism regulation. Regarding proteomics, top-down proteomics is an approach which is growing within this field due to its capacity to identify many PTMs in complex proteomes. The objective of this project was to characterize the proteome of C. glutamicum, mainly those proteins related to amino acids metabolism. Also, propose regulations of mechanisms that can enhance amino acid production in this bacterium. Top-down proteomics experiments were performed in C. glutamicum samples cultivated in BT medium and CGXII medium. In BT medium were detected 1873 PrSMs having identified 492 were non-redundant proteins with 681 proteoforms. Those proteoforms were related to cleaveges and addition of groups such as formyl, acetyl and phosphates. These analyses were also able to detect sample proteolysis degradation through spectral counting. The CGXII medium was submitted to different conditions: i) with glutamate production, induced by addition of tween 40 and, ii) control condition, without tween 40. Glutamate production was observed after 30 hs of cultivation with tween 40 in contrast to the sample without this molecule, where glutamate production was not detected. Despite this phenotype, no significant changes in modifications patterns was observed by means of spectral counting. Furthermore, in this sample, the degradation effect observed was much less than the one detected in BT medium samples. Phosphorylated proteoforms identified suggest a potential amino acid metabolism regulation. Such proteins as lactate dehydrogenase and threonine dehydratase detected with phosphate addition group might infer a regulation in the biosynthesis of L-glutamate, L-lysine and L-Leucine. Moreover, it was identified mass shift patterns, suggesting different amounts of the same PTM into two proteins (3-isopropylmalate dehydratase and acetolactate synthase, annotated by similarity). These two proteins catalyze biosynthesis steps of L-valine and L-leucine. The mass shifts present in theses suggest different levels of stearoylation modification, possibly a new mechanism of regulation in this amino acid synthesis. Beyond group additions, proteoforms were detected with small sequence cleavages. Among them, chorismate mutase drives special attention because this protein is part of a supercomplex formed by chorismate mutase and DAHP synthase. Consequently, this cleavage could disturb the production of L-phenylalanine and L-tyrosine.
metadata.dc.description.unidade:	Instituto de Ciências Biológicas (IB) Departamento de Biologia Celular (IB CEL)
Description:	Dissertação (mestrado)—Universidade de Brasília, Instituto de Biologia, Pós-Graduação em Biologia Microbiana, 2019.
metadata.dc.description.ppg:	Programa de Pós-Graduação em Biologia Microbiana
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