Artigo em elaboração _Reconstrucao de redes metabólicas

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ORIGINAL RESEARCH PROPOSAL
Genomic-scale reconstruction of the metabolic network in
Burkholderia sacchari to expand its biotechnological applications
ABSTRACT
Burkholderia sacchari have demonstrated the ability to produce different bioproducts. To expand the knowledge
of its metabolism, in order to expand its biotechnological applications, knowledge of its metabolic network
becomes essential. In this work, the reconstruction of the genomic-scale metabolic network of B sacchari was
developed based on multi-omics analyses. Genome general information indicates 7,317,626 base pairs (bp), a
GC content of 64%, 51 paralogous genes and 6,975 coding regions. After manual curing and prediction, there
was a wide distribution in the genome of genes related to major superfamily transporters (MFS) close to genes
involved in the metabolism of xylose, galactose, arabinose, glucose, and aromatic compounds. The influence of
these transporters on the metabolization of these carbon sources was studied. The organization of glycerol
consumption genes was explored, since this substrate is frequently used in industry, and its organization differs
from the organization in Pseudomonas putida, a microorganism efficient in metabolizing this substrate. The
ability to produce metabolites such as Methylglyoxal and L-Ribose were also considered, since they have
properties of biotechnological interest. The analysis indicated the partial presence of the genetic apparatus for
the synthesis of these molecules. Regarding PHA, a bioproduct candidate to replace conventional plastics, two
class I PHA synthases were identified, however, only one is structurally conserved. The phylogenetic
relationship was explored in order to search for reference strains, indicating 98.28% proximity to
Paraburkholderia dokdonella. Approaches to assess and scale up genome quality were applied, resulting in a
reduction in the number of contigs from 21 to 5, and alignment from 9.5% to 60%. Aspects related to biosafety
were raised through the analysis of resistance to antibiotics and virulence factors, indicating that it is a safe
strain. The first draft of the metabolic model indicates the presence of the Entner-Doudoroff (ED) pathway and
the absence of the Embden-Meyerhof-Parnas (EMP) pathway and 127 reactions, 144 metabolites and 246 genes.
In the Krebs cycle (CK), the enzyme malate quinone oxidoreductase, responsible for converting oxaloacetate to
malate, is absent, this reaction is carried out by malate dehydrogenase. The dependency on NAD or NADP
cofactors of three enzymes was investigated in order to refine the metabolic model. In a bioreactor, experiments
were carried out providing glucose, fructose, sucrose, fructose + glucose, xylose and glycerol. These assays
were used in fluxomic analysis since they allowed to delimit the maximum specific growth velocity and the
carbon source conversion efficiency. The metabolic model reconstructed here represents a unique tool to predict
success in the production of different bioproducts performed by different strains of Burkholderia sacchari.
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