Edson L. Folador

Proteome scale comparative modeling for conserved drug and vaccine targets identification in Corynebacterium pseudotuberculosis

Corynebacterium pseudotuberculosis (Cp) is a pathogenic bacterium that causes caseous lymphadenitis (CLA), ulcerative lymphangitis, mastitis, and edematous to a broad spectrum of hosts, including ruminants, thereby threatening economic and dairy industries worldwide. Currently there is no effective drug or vaccine available against Cp. To identify new targets, we adopted a novel integrative strategy, which began with the prediction of the modelome (tridimensional protein structures for the proteome of an organism, generated through comparative modeling) for 15 previously sequenced C. pseudotuberculosis strains. This pan-modelomics approach identified a set of 331 conserved proteins having 95-100% intra-species sequence similarity. Next, we combined subtractive proteomics and modelomics to reveal a set of 10 Cp proteins, which may be essential for the bacteria. Of these, 4 proteins (tcsR, mtrA, nrdI, and ispH) were essential and non-host homologs (considering man, horse, cow and sheep as hosts) and satisfied all criteria of being putative targets. Additionally, we subjected these 4 proteins to virtual screening of a drug-like compound library. In all cases, molecules predicted to form favorable interactions and which showed high complementarity to the target were found among the top ranking compounds. The remaining 6 essential proteins (adk, gapA, glyA, fumC, gnd, and aspA) have homologs in the host proteomes. Their active site cavities were compared to the respective cavities in host proteins. We propose that some of these proteins can be selectively targeted using structure-based drug design approaches (SBDD). Our results facilitate the selection of C. pseudotuberculosis putative proteins for developing broad-spectrum novel drugs and vaccines. A few of the targets identified here have been validated in other microorganisms, suggesting that our modelome strategy is effective and can also be applicable to other pathogens.

Genome Sequence of Lactococcus lactis subsp. lactis NCDO 2118, a GABA-Producing Strain

ABSTRACT Lactococcus lactis subsp. lactis NCDO 2118 is a nondairy lactic acid bacterium, a xylose fermenter, and a gamma-aminobutyric acid (GABA) producer isolated from frozen peas. Here, we report the complete genome sequence of L. lactis NCDO 2118, a strain with probiotic potential activity.

Label-free proteomic analysis to confirm the predicted proteome of Corynebacterium pseudotuberculosis under nitrosative stress mediated by nitric oxide

BackgroundCorynebacterium pseudotuberculosis biovar ovis is a facultative intracellular pathogen, and the etiological agent of caseous lymphadenitis in small ruminants. During the infection process, the bacterium is subjected to several stress conditions, including nitrosative stress, which is caused by nitric oxide (NO). In silico analysis of the genome of C. pseudotuberculosis ovis 1002 predicted several genes that could influence the resistance of this pathogen to nitrosative stress. Here, we applied high-throughput proteomics using high definition mass spectrometry to characterize the functional genome of C. pseudotuberculosis ovis 1002 in the presence of NO-donor Diethylenetriamine/nitric oxide adduct (DETA/NO), with the aim of identifying proteins involved in nitrosative stress resistance.ResultsWe characterized 835 proteins, representing approximately 41% of the predicted proteome of C. pseudotuberculosis ovis 1002, following exposure to nitrosative stress. In total, 102 proteins were exclusive to the proteome of DETA/NO-induced cells, and a further 58 proteins were differentially regulated between the DETA/NO and control conditions. An interactomic analysis of the differential proteome of C. pseudotuberculosis in response to nitrosative stress was also performed. Our proteomic data set suggested the activation of both a general stress response and a specific nitrosative stress response, as well as changes in proteins involved in cellular metabolism, detoxification, transcriptional regulation, and DNA synthesis and repair.ConclusionsOur proteomic analysis validated previously-determined in silico data for C. pseudotuberculosis ovis 1002. In addition, proteomic screening performed in the presence of NO enabled the identification of a set of factors that can influence the resistance and survival of C. pseudotuberculosis during exposure to nitrosative stress.

In silico identification of essential proteins in Corynebacterium pseudotuberculosis based on protein-protein interaction networks.

BackgroundCorynebacterium pseudotuberculosis (Cp) is a gram-positive bacterium that is classified into equi and ovis serovars. The serovar ovis is the etiological agent of caseous lymphadenitis, a chronic infection affecting sheep and goats, causing economic losses due to carcass condemnation and decreased production of meat, wool, and milk. Current diagnosis or treatment protocols are not fully effective and, thus, require further research of Cp pathogenesis.ResultsHere, we mapped known protein-protein interactions (PPI) from various species to nine Cp strains to reconstruct parts of the potential Cp interactome and to identify potentially essential proteins serving as putative drug targets. On average, we predict 16,669 interactions for each of the nine strains (with 15,495 interactions shared among all strains). An in silico sanity check suggests that the potential networks were not formed by spurious interactions but have a strong biological bias. With the inferred Cp networks we identify 181 essential proteins, among which 41 are non-host homologous.ConclusionsThe list of candidate interactions of the Cp strains lay the basis for developing novel hypotheses and designing according wet-lab studies. The non-host homologous essential proteins are attractive targets for therapeutic and diagnostic proposes. They allow for searching of small molecule inhibitors of binding interactions enabling modern drug discovery. Overall, the predicted Cp PPI networks form a valuable and versatile tool for researchers interested in Corynebacterium pseudotuberculosis.

Genome Sequence of Corynebacterium pseudotuberculosis MB20 bv. equi Isolated from a Pectoral Abscess of an Oldenburg Horse in California.

ABSTRACT The genome of Corynebacterium pseudotuberculosis MB20 bv. equi was sequenced using the Ion Personal Genome Machine (PGM) platform, and showed a size of 2,363,089 bp, with 2,365 coding sequences and a GC content of 52.1%. These results will serve as a basis for further studies on the pathogenicity of C. pseudotuberculosis bv. equi.

Label-free quantitative proteomics of Corynebacterium pseudotuberculosis isolates reveals differences between Biovars ovis and equi strains

BackgroundCorynebacterium pseudotuberculosis is a pathogen classified into two biovars: C. pseudotuberculosis biovar ovis, the etiologic agent of caseous lymphadenitis and C. pseudotuberculosis biovar equi, which causes ulcerative lymphangitis. The available whole genome sequences of different C. pseudotuberculosis strains have enabled identify difference of genes related both virulence and physiology of each biovar. To evaluate be this difference could reflect at proteomic level and to better understand the shared factors and the exclusive ones of biovar ovis and biovar equi strains, we applied the label-free quantitative proteomic to characterize the proteome of the strains: 1002_ovis and 258_equi, isolated from goat (Brazil) and equine (Belgium), respectively.ResultsFrom this analysis, we characterized a total of 1230 proteins in 1002_ovis and 1220 in 258_equi with high confidence. Moreover, the core-proteome between 1002_ovis and 258_equi obtained here is composed of 1122 proteins involved in different cellular processes, which could be necessary for the free living of C. pseudotuberculosis. In addition, 120 proteins from this core-proteome presented change in abundant with statistically significant differences. Considering the exclusive proteome, we detected strain-specific proteins to each strain. When correlated, the exclusive proteome of each strain and proteome with change in abundant, the proteomic differences, between the 1002_ovis and 258_equi, this related to proteins involved in cellular metabolism, information storage and processing, cellular processes and signaling.ConclusionsThis study reports the first comparative proteomic study of the biovars ovis and equi of C. pseudotuberculosis. The results generated in this study provide information about factors which can contribute to understanding both the physiology and the virulence of this pathogen.

Genome Sequence of Corynebacterium ulcerans Strain 210932

ABSTRACT In this work, we present the complete genome sequence of Corynebacterium ulcerans strain 210932, isolated from a human. The species is an emergent pathogen that infects a variety of wild and domesticated animals and humans. It is associated with a growing number of cases of a diphtheria-like disease around the world.

SIMBA: a web tool for managing bacterial genome assembly generated by Ion PGM sequencing technology

BackgroundThe evolution of Next-Generation Sequencing (NGS) has considerably reduced the cost per sequenced-base, allowing a significant rise of sequencing projects, mainly in prokaryotes. However, the range of available NGS platforms requires different strategies and software to correctly assemble genomes. Different strategies are necessary to properly complete an assembly project, in addition to the installation or modification of various software. This requires users to have significant expertise in these software and command line scripting experience on Unix platforms, besides possessing the basic expertise on methodologies and techniques for genome assembly. These difficulties often delay the complete genome assembly projects.ResultsIn order to overcome this, we developed SIMBA (SImple Manager for Bacterial Assemblies), a freely available web tool that integrates several component tools for assembling and finishing bacterial genomes. SIMBA provides a friendly and intuitive user interface so bioinformaticians, even with low computational expertise, can work under a centralized administrative control system of assemblies managed by the assembly center head. SIMBA guides the users to execute assembly process through simple and interactive pages. SIMBA workflow was divided in three modules: (i) projects: allows a general vision of genome sequencing projects, in addition to data quality analysis and data format conversions; (ii) assemblies: allows de novo assemblies with the software Mira, Minia, Newbler and SPAdes, also assembly quality validations using QUAST software; and (iii) curation: presents methods to finishing assemblies through tools for scaffolding contigs and close gaps. We also presented a case study that validated the efficacy of SIMBA to manage bacterial assemblies projects sequenced using Ion Torrent PGM.ConclusionBesides to be a web tool for genome assembly, SIMBA is a complete genome assemblies project management system, which can be useful for managing of several projects in laboratories. SIMBA source code is available to download and install in local webservers at http://ufmg-simba.sourceforge.net.

Genome Sequence of Corynebacterium ulcerans Strain FRC11

ABSTRACT Here, we present the genome sequence of Corynebacterium ulcerans strain FRC11. The genome includes one circular chromosome of 2,442,826 bp (53.35% G+C content), and 2,210 genes were predicted, 2,146 of which are putative protein-coding genes, with 12 rRNAs and 51 tRNAs; 1 pseudogene was also identified.

Comparative proteomic analysis of four biotechnological strains Lactococcus lactis through label-free quantitative proteomics

Lactococcus lactis is a bacteria with high biotechnological potential, where is frequently used in the amino acid production and production of fermented dairy products, as well as drug delivery systems and mucosal vaccine vector. The knowledge of a functional core proteome is important extremely for both fundamental understanding of cell functions and for synthetic biology applications. In this study, we characterized the L. lacits proteome from proteomic analysis of four biotechnological strains L. lactis: L. lactis subsp. lactis NCDO2118, L. lactis subsp. lactis IL1403, L. lactis subsp. cremoris NZ9000 and L. lactis subsp. cremoris MG1363. Our label‐free quantitative proteomic analysis of the whole bacterial lysates from each strains resulted in the characterization of the L. lactis core proteome that was composed by 586 proteins, which might contribute to resistance of this bacterium to different stress conditions as well as involved in the probiotic characteristic of L. lactis. Kegg enrichment analysis shows that ribosome, metabolic pathways, pyruvate metabolism and microbial metabolism in diverse environments were the most enriched. According to our quantitative proteomic analysis, proteins related to translation process were the more abundant in the core proteome, which represent an important step in the synthetic biology. In addition, we identified a subset of conserved proteins that are exclusive of the L. lactis subsp. cremoris or L. lactis subsp. lactis, which some are related to metabolic pathway exclusive. Regarding specific proteome of NCDO2118, we detected ‘strain‐specific proteins’. Finally, proteogenomics analysis allows the identification of proteins, which were not previously annotated in IL1403 and MG1363. The results obtained in this study allowed to increase our knowledge about the biology of L. lactis, which contributes to the implementation of strategies that make it possible to increase the biotechnological potential of this bacterium.