Carbohydrate metabolism of Xylella fastidiosa: Detection of glycolytic and pentose phosphate pathway enzymes and cloning and expression of the enolase gene

Facincani, Agda Paula; Ferro, Jesus Aparecido; Pizauro, João Martins; Pereira, H. A.; Lemos, Eliana Gertrudes de Macedo; Prado, Alessandro Luis do; Ferro, Maria Inês Tiraboschi

doi:10.1590/s1415-47572003000200015

Cited by 5 publications

(5 citation statements)

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“…Therefore, the incorrect folding of the protein HpEno should not be the most plausible cause for the lack of activity. Some reports indicated that enolases from X. fastidiosa and chicken muscle and liver were irreversibly denatured by urea, suggesting that this chaotropic agent should not be used to extract enolase from inclusion bodies [57]. In contrast, we have found 85-90% reversibility of the urea-induced unfolding reaction for yeast enolase [58].…”

Section: Discussionmentioning

confidence: 75%

Biochemical and Biophysical Characterization of the Enolase fromHelicobacter pylori

López-López

Rodríguez-Luna

Lara-Ramírez

et al. 2018

BioMed Research International

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Enolase, which catalyses the conversion of 2-phospho-D-glycerate to phosphoenolpyruvate, is an important enzyme in the classic glycolysis pathway in cells. Enolase is highly conserved in organisms from bacteria to humans, indicating its importance in cells. Thus, enolase is a good target for developing new drugs. In the last decade, new functions of this enzyme have been found. Helicobacter pylori is a common human pathogen that causes gastric diseases and even gastric cancer. In this study, the sequence of H. pylori enolase (HpEno) was analysed; the conservation (at least partial) of binding sites for cofactor, plasminogen, and host extracellular RNA, as well as catalytic site, indicates that HpEno should be capable of performing the functions. Recombinant HpEno was overexpressed and purified from E. coli. Compared to the enolases from other species, HpEno had similar characteristics for its secondary structure. The temperature-induced profiles indicate that HpEno is quite stable to temperature, compared to other homologs. Regarding the kinetics of the unfolding reaction, we found that the activation enthalpy associated with the thermal unfolding reaction is equivalent to the reported activation enthalpy for yeast enolase, indicating a similar scaffold and kinetic stability. Although a wide range of experimental conditions were assayed, it was not possible to detect any enzymatic activity of HpEno. To prove the lack of activity, still a much wider range of experiments should be carried out.

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Section: Discussionmentioning

confidence: 75%

Biochemical and Biophysical Characterization of the Enolase fromHelicobacter pylori

López-López

Rodríguez-Luna

Lara-Ramírez

et al. 2018

BioMed Research International

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“…Several xanthomonads use the Entner-Doudoroff pathway (EMP) as the main catabolic route for glucose [50], including X. campestris [38]. Although this was not demonstrated for X. fastidiosa yet, due to the genetic and metabolic similarities among these organisms, the EMP has been proposed to be the main glucose degradation pathway for X. fastidiosa [51].…”

Section: Resultsmentioning

confidence: 99%

An updated genome-scale model forXylella fastidiosasubsp.paucaDe Donno

Oliveira

Cunha

Silva

et al. 2022

Preprint

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Xylella fastidiosa is a gram-negative phytopathogenic bacterium that caused a significant economic impact around the world. In the last decade, genome-scale metabolic models have become important systems biology tools for studying the metabolic behaviour of different pathogens and driving the discovery of novel drug targets. In this work, a GSM model of X. fastidiosa subsp. pauca De Donno was developed. The model comprises 1164 reactions, 1379 metabolites, and 508 genes. in silico validation of the metabolic model was achieved through the comparison of simulations with available experimental data. Aerobic metabolism was simulated properly and fastidian gum production rates predicted accurately. The GSM model allowed identifying potential drug targets, using a pipeline based on the model's gene essentially analysis.

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“…The functionality of the glycolytic pathway in X. fastidiosa was evaluated ( Facincani et al , 2003 ) by studying the enzymes phosphoglucose isomerase, aldolase, glyceraldehyde-3-phosphate dehydrogenase and pyruvate kinase from the glycolytic pathway, and glucose-6-phosphate dehydrogenase from the Entner-Doudoroff, followed by cloning and expression studies of the enolase gene and determination of its activity. These studies showed that X. fastidiosa does not use the glycolytic pathway to metabolize carbohydrates.…”

Section: Resultsmentioning

confidence: 99%

Xylella fastidiosa gene expression analysis by DNA microarrays

et al. 2009

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Xylella fastidiosa genome sequencing has generated valuable data by identifying genes acting either on metabolic pathways or in associated pathogenicity and virulence. Based on available information on these genes, new strategies for studying their expression patterns, such as microarray technology, were employed. A total of 2,600 primer pairs were synthesized and then used to generate fragments using the PCR technique. The arrays were hybridized against cDNAs labeled during reverse transcription reactions and which were obtained from bacteria grown under two different conditions (liquid XDM 2 and liquid BCYE). All data were statistically analyzed to verify which genes were differentially expressed. In addition to exploring conditions for X. fastidiosa genome-wide transcriptome analysis, the present work observed the differential expression of several classes of genes (energy, protein, amino acid and nucleotide metabolism, transport, degradation of substances, toxins and hypothetical proteins, among others). The understanding of expressed genes in these two different media will be useful in comprehending the metabolic characteristics of X. fastidiosa, and in evaluating how important certain genes are for the functioning and survival of these bacteria in plants.

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Carbohydrate metabolism of Xylella fastidiosa: Detection of glycolytic and pentose phosphate pathway enzymes and cloning and expression of the enolase gene

Cited by 5 publications

References 16 publications

Biochemical and Biophysical Characterization of the Enolase fromHelicobacter pylori

Biochemical and Biophysical Characterization of the Enolase fromHelicobacter pylori

An updated genome-scale model forXylella fastidiosasubsp.paucaDe Donno

Xylella fastidiosa gene expression analysis by DNA microarrays

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