scholarly journals Pathways Analyzer: Design of a Tool for the Synthetic Assembly of Escherichia Coli K-12 MG1655 Bacteria for Biofuel Production

2019 ◽  
Author(s):  
Laura Vasquez ◽  
Ricardo Alvarado ◽  
Allan Orozco
Keyword(s):  
Synthetic Biology ◽  
Metabolic Pathways ◽  
Point Of View ◽  
Biofuel Production ◽  
Design Stage ◽  
Computational Point ◽  
E Coli ◽  
Process Waste ◽  
K 12 ◽  
The Impact

ABSTRACTSummaryDue to the impact of environmental pollution, the importance of producing high quality biofuels and to leverage organic waste that normally would have no use has increased over time. Through synthetic biology, it is possible to improve existing organisms to process waste that is traditionally not used for biofuel production, such as whey.With the redesign of metabolic pathways, it is possible to create connections for the implementation of new organisms that carry out functions that are normally not present in nature.From a computational point of view, metabolic pathways, which can be found in data sources as KEGG, can be converted to a graph data structure. These transformations enable the use of well-known algorithms, which enables the optimization of the analyses required to achieve the assembly of new organisms.The present work aims to design a tool for the transformation of metabolic pathways and the development of path finding algorithms that establish relevant links between compounds that are essential to the biofuel production process.As a result, a catalog of biobricks is created from the analysis of a subset of paths which can be used in the design stage of the synthetic assembly of the E. coli bacteria. The assembly’s structure and functions are characterized according to the pieces used.Finally, new constructions are visualized with the goal of demonstrating and supporting the analysis processes, thus assisting people that work in the field of Synthetic Biology.AvailabilityPathways Analyzer is accessible at: https://gitlab.com/lvasquezcr/pathways-analyzer/

scholarly journals Role of Escherichia coli O157:H7 Virulence Factors in Colonization at the Bovine Terminal Rectal Mucosa

2006 ◽  
Vol 74 (8) ◽  
pp. 4685-4693 ◽  
Author(s):  
Haiqing Sheng ◽  
Ji Youn Lim ◽  
Hannah J. Knecht ◽  
Jie Li ◽  
Carolyn J. Hovde
Keyword(s):  
Escherichia Coli ◽  
Virulence Factors ◽  
Clinical Manifestations ◽  
Rectal Mucosa ◽  
Wild Type ◽  
E Coli ◽  
Healthy Cattle ◽  
Life Threatening ◽  
K 12 ◽  
The Impact

ABSTRACT The human pathogen Escherichia coli O157:H7 causes hemorrhagic colitis and life-threatening sequelae and transiently colonizes healthy cattle at the terminal rectal mucosa. This study analyzed virulence factors important for the clinical manifestations of human E. coli O157:H7 infection for their contribution to the persistence of E. coli in cattle. The colonizing ability of E. coli O157:H7 was compared with those of nonpathogenic E. coli K-12 and isogenic deletion mutants missing Shiga toxin (Stx), the adhesin intimin, its receptor Tir, hemolysin, or the ∼92-kb pO157. Fully ruminant steers received a single rectal application of one E. coli strain so that effects of mucosal attachment and survival at the terminal rectum could be measured without the impact of bacterial passage through the entire gastrointestinal tract. Colonization was monitored by sensitive recto-anal junction mucosal swab culture. Nonpathogenic E. coli K-12 did not colonize as well as E. coli O157:H7 at the bovine terminal rectal mucosa. The E. coli O157:H7 best able to persist had intimin, Tir, and the pO157. Strains missing even one of these factors were recovered in lower numbers and were cleared faster than the wild type. In contrast, E. coli O157:H7 strains that were missing Stx or hemolysin colonized like the wild type. For these three strains, the number of bacteria increased between days 1 and 4 postapplication and then decreased slowly. In contrast, the numbers of noncolonizing strains (K-12, Δtir, and Δeae) decreased from the day of application. These patterns consistently predicted long-term colonization or clearance of the bacteria from the bovine terminal rectal mucosa.

scholarly journals Robustness testing and scalability of phosphate regulated promoters useful for two-stage autoinduction in E. coli

2020 ◽  
Author(s):  
Eirik A Moreb ◽  
Zhixia Ye ◽  
John P Efromson ◽  
Jennifer N Hennigan ◽  
Romel Menacho-Melgar ◽  
...  
Keyword(s):  
Synthetic Biology ◽  
Heterologous Protein ◽  
Scale Up ◽  
Mineral Salts ◽  
Two Stage ◽  
Robustness Testing ◽  
E Coli ◽  
Media Formulation ◽  
The Impact ◽  
Regulated Promoters

A key challenge in synthetic biology is the successful utilization of characterized parts, such as promoters, in different biological contexts. We report the robustness testing of a small library of E. coli PhoB regulated promoters that enable heterologous protein production in two-stage cultures. Expression levels were measured both in a rich Autoinduction Broth as well as a minimal mineral salts media. Media dependent differences were promoter dependent. 4 out of 16 promoters tested were identified to have tightly controlled expression which was also robust to media formulation. Improved promoter robustness led to more predictable scale up and consistent expression in instrumented bioreactors. This subset of PhoB activated promoters, useful for two-stage autoinduction, highlight the impact of the environment on the performance of biological parts, and the importance of robustness testing in synthetic biology.

Punching Force of Cold Die of Fuzzy Reliability Considering the Random Distribution of Material Properties

2011 ◽  
Vol 311-313 ◽  
pp. 1577-1582
Author(s):  
Zhi Su Zhao ◽  
Xing Hua Zhang
Keyword(s):  
Material Properties ◽  
Random Distribution ◽  
Point Of View ◽  
Design Stage ◽  
Gradual Change ◽  
Fuzzy Reliability ◽  
Reliability Design ◽  
Curve Design ◽  
Processing Error ◽  
The Impact

In order to be considered in the calculation of punching force of cold die random distribution of material properties, meanwhile including success and failure punching during the gradual change process in the calculation. Based on probabilistic fuzzy reliability point of view, the success or failure determination will be extended to fuzzy events. Based on fuzzy reliability, the punching force of cold die calculation method will be also given when taking the impact of random distribution of material properties into account. Related analysis formulae and the fuzzy criterion of success or failure of punching are established and derived. Through which, design and engineering process are integrated, the designer will be more reliably to predict the success or failure of the punching during the design stage. The processing error of lack of statistical data and the objectivity of the success or failure determination criterion will be easily solved. Economy cost and reliability design of geometrical curve design will be also considered.

scholarly journals Smart Cooperative Energy Supply Strategy to Increase Reliability in Residential Stand-Alone Photovoltaic Systems

2021 ◽  
Vol 11 (24) ◽  
pp. 11723
Author(s):  
Carlos Roldán-Blay ◽  
Carlos Roldán-Porta ◽  
Eduardo Quiles ◽  
Guillermo Escrivá-Escrivá
Keyword(s):  
Rural Areas ◽  
Energy Supply ◽  
Sequential Monte Carlo ◽  
Residential Buildings ◽  
Cost Effective ◽  
Point Of View ◽  
Design Stage ◽  
Interconnected System ◽  
Pv Systems ◽  
The Impact

In reliability studies of isolated energy supply systems for residential buildings, supply failures due to insufficient generation are generally analysed. Recent studies conclude that this kind of analysis makes it possible to optimally design the sizes of the elements of the generation system. However, in isolated communities or rural areas, it is common to find groups of dwellings in which micro-renewable sources, such as photovoltaic (PV) systems, can be installed. In this situation, the generation and storage of several houses can be considered as an interconnected system forming a cooperative microgrid (CoMG). This work analyses the benefits that sharing two autonomous installations can bring to each one, from the point of view of reliability. The method consists of the application of a random sequential Monte Carlo (SMC) simulation to the CoMG to evaluate the impact of a simple cooperative strategy on the reliability of the set. The study considers random failures in the generation systems. The results show that the reliability of the system increases when cooperation is allowed. Additionally, at the design stage, this allows more cost-effective solutions than single sizing with a similar level of reliability.

scholarly journals Peptide Transporter CstA Imports Pyruvate in Escherichia coli K-12

2018 ◽  
Vol 200 (7) ◽  
Author(s):  
Soonkyu Hwang ◽  
Donghui Choe ◽  
Minseob Yoo ◽  
Sanghyuk Cho ◽  
Sun Chang Kim ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
High Throughput ◽  
Metabolic Pathways ◽  
Genetic Information ◽  
Peptide Transporter ◽  
Pyruvate Metabolism ◽  
Content Type ◽  
E Coli ◽  
Uptake Activity ◽  
K 12

ABSTRACT Pyruvate is an important intermediate of central carbon metabolism and connects a variety of metabolic pathways in Escherichia coli . Although the intracellular pyruvate concentration is dynamically altered and tightly balanced during cell growth, the pyruvate transport system remains unclear. Here, we identified a pyruvate transporter in E. coli using high-throughput transposon sequencing. The transposon mutant library (a total of 5 × 10 5 mutants) was serially grown with a toxic pyruvate analog (3-fluoropyruvate [3FP]) to enrich for transposon mutants lacking pyruvate transport function. A total of 52 candidates were selected on the basis of a stringent enrichment level of transposon insertion frequency in response to 3FP treatment. Subsequently, their pyruvate transporter function was examined by conventional functional assays, such as those measuring growth inhibition by the toxic pyruvate analog and pyruvate uptake activity. The pyruvate transporter system comprises CstA and YbdD, which are known as a peptide transporter and a conserved protein, respectively, whose functions are associated with carbon starvation conditions. In addition to the presence of more than one endogenous pyruvate importer, it has been suggested that the E. coli genome encodes constitutive and inducible pyruvate transporters. Our results demonstrated that CstA and YbdD comprise the constitutive pyruvate transporter system in E. coli , which is consistent with the tentative genomic locus previously suggested and the functional relationship with the extracellular pyruvate sensing system. The identification of this pyruvate transporter system provides valuable genetic information for understanding the complex process of pyruvate metabolism in E. coli . IMPORTANCE Pyruvate is an important metabolite as a central node in bacterial metabolism, and its intracellular levels are tightly regulated to maintain its functional roles in highly interconnected metabolic pathways. However, an understanding of the mechanism of how bacterial cells excrete and transport pyruvate remains elusive. Using high-throughput transposon sequencing followed by pyruvate uptake activity testing of the selected candidate genes, we found that a pyruvate transporter system comprising CstA and YbdD, currently annotated as a peptide transporter and a conserved protein, respectively, constitutively transports pyruvate. The identification of the physiological role of the pyruvate transporter system provides valuable genetic information for understanding the complex pyruvate metabolism in Escherichia coli .

Geometric Curve Design Method of Fuzzy Reliability Based on Random Processing Errors

2011 ◽  
Vol 199-200 ◽  
pp. 72-77
Author(s):  
Zhi Su Zhao ◽  
Xing Hua Zhang
Keyword(s):  
Design Method ◽  
Failure Process ◽  
Point Of View ◽  
Design Stage ◽  
Gradual Change ◽  
Fuzzy Reliability ◽  
Reliability Design ◽  
Curve Design ◽  
Processing Errors ◽  
The Impact

In order to foresee the influence of random processing errors on geometric curve in design stage, meanwhile including success and failure process during the gradual change process in the forecast. Based on probabilistic fuzzy reliability point of view, the success or failure determination will be extended to fuzzy events. The geometric curve deign method will be also given when taking the impact of random engineering error into account. Related analysis formulae and the fuzzy criterion of success or failure of designing the curve process are established and derived. Through which, design and engineering process are integrated, the designer will be more reliably to predict the success or failure of the geometric curve design during the design stage. The processing error of lack of statistical data and the objectivity of the success or failure determination criterion will be easily solved. Economy cost and reliability design of geometrical curve design will be also considered.

scholarly journals Model System To Evaluate the Effect of ampD Mutations on AmpC-Mediated β-Lactam Resistance

2006 ◽  
Vol 50 (6) ◽  
pp. 2030-2037 ◽  
Author(s):  
Amber J. Schmidtke ◽  
Nancy D. Hanson
Keyword(s):  
Escherichia Coli ◽  
Pseudomonas Aeruginosa ◽  
Structural Gene ◽  
Model System ◽  
Alternative Mechanism ◽  
Citrobacter Freundii ◽  
E Coli ◽  
K 12 ◽  
Carboxy Terminal ◽  
The Impact

ABSTRACT Mutations within the structural gene of ampD can lead to AmpC overproduction and increases in β-lactam MICs in organisms with an inducible ampC. However, identification of mutations alone cannot predict the impact that those mutations have on AmpD function. Therefore, a model system was designed to determine the effect of ampD mutations on ceftazidime MICs using an AmpD− mutant Escherichia coli strain which produced an inducible plasmid-encoded AmpC. ampD genes were amplified by PCR from strains of E. coli, Citrobacter freundii, and Pseudomonas aeruginosa. Also, carboxy-terminal truncations of C. freundii ampD genes were constructed representing deletions of 10, 21, or 25 codons. Amplified ampD products were cloned into pACYC184 containing inducible bla ACT-1-ampR. Plasmids were transformed into E. coli strains JRG582 (AmpD−) and K-12 259 (AmpD+). The strains were evaluated for a derepressed phenotype using ceftazidime MICs. Some mutated ampD genes, including the ampD gene of a derepressed C. freundii isolate, resulted in substantial decreases in ceftazidime MICs (from >256 μg/ml to 12 to 24 μg/ml) for the AmpD− strain, indicating no role for these mutations in derepressed phenotypes. However, ampD truncation products and ampD from a partially derepressed P. aeruginosa strain resulted in ceftazidime MICs of >256 μg/ml, indicating a role for these gene modifications in derepressed phenotypes. The use of this model system indicated that alternative mechanisms were involved in the derepressed phenotype observed in strains of C. freundii and P. aeruginosa. The alternative mechanism involved in the derepressed phenotype of the C. freundii isolate was downregulation of ampD transcription.

scholarly journals Indole-3-acetic acid regulates the central metabolic pathways in Escherichia coli

Microbiology ◽  
2006 ◽  
Vol 152 (8) ◽  
pp. 2421-2431 ◽  
Author(s):  
C. Bianco ◽  
E. Imperlini ◽  
R. Calogero ◽  
B. Senatore ◽  
P. Pucci ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Metabolic Pathways ◽  
Tricarboxylic Acid Cycle ◽  
Tricarboxylic Acid ◽  
Mrna Levels ◽  
Glyoxylate Shunt ◽  
Acetyl Coa ◽  
Acid Cycle ◽  
E Coli ◽  
K 12

The physiological changes induced by indoleacetic acid (IAA) treatment were investigated in the totally sequenced Escherichia coli K-12 MG1655. DNA macroarrays were used to measure the mRNA levels for all the 4290 E. coli protein-coding genes; 50 genes (1.1 %) exhibited significantly different expression profiles. In particular, genes involved in the tricarboxylic acid cycle, the glyoxylate shunt and amino acid biosynthesis (leucine, isoleucine, valine and proline) were up-regulated, whereas the fermentative adhE gene was down-regulated. To confirm the indications obtained from the macroarray analysis the activity of 34 enzymes involved in central metabolism was measured; this showed an activation of the tricarboxylic acid cycle and the glyoxylate shunt. The malic enzyme, involved in the production of pyruvate, and pyruvate dehydrogenase, required for the channelling of pyruvate into acetyl-CoA, were also induced in IAA-treated cells. Moreover, it was shown that the enhanced production of acetyl-CoA and the decrease of NADH/NAD+ ratio are connected with the molecular process of the IAA response. The results demonstrate that IAA treatment is a stimulus capable of inducing changes in gene expression, enzyme activity and metabolite level involved in central metabolic pathways in E. coli.

scholarly journals Phenotype prediction in anEscherichia colistrain panel

2017 ◽  
Author(s):  
Marco Galardin ◽  
Alexandra Koumoutsi ◽  
Lucia Herrera-Dominguez ◽  
Juan Antonio Cordero Varela ◽  
Anja Telzerow ◽  
...  
Keyword(s):  
High Throughput ◽  
Gene Function ◽  
Genetic Variants ◽  
List Type ◽  
Loss Of Function ◽  
Phenotypic Differences ◽  
E Coli ◽  
Growth Defects ◽  
K 12 ◽  
The Impact

SummaryUnderstanding how genetic variation contributes to phenotypic differences is a fundamental question in biology. Here, we set to predict fitness defects of an individual using mechanistic models of the impact of genetic variants combined with prior knowledge of gene function. We assembled a diverse panel of 696Escherichia colistrains for which we obtained genomes and measured growth phenotypes in 214 conditions. We integrated variant effect predictors to derive gene-level probabilities of loss of function for every gene across strains. We combined these probabilities with information on conditional gene essentiality in the reference K-12 strain to predict the strains’ growth defects, providing significant predictions for up to 38% of tested conditions. The putative causal variants were validated in complementation assays highlighting commonly perturbed pathways in evolution for the emergence of growth phenotypes. Altogether, our work illustrates the power of integrating high-throughput gene function assays to predict the phenotypes of individuals.HighlightsAssembled a reference panel ofE. colistrainsGenotyped and high-throughput phenotyped theE. colireference strain panelReliably predicted the impact of genetic variants in up to 38% of tested conditionsHighlighted common genetic pathways for the emergence of deleterious phenotypes

scholarly journals Unexpected Phytostimulatory Behavior for Escherichia coli and Agrobacterium tumefaciens Model Strains

2013 ◽  
Vol 26 (5) ◽  
pp. 495-502 ◽  
Author(s):  
Vincent Walker ◽  
Maxime Bruto ◽  
Floriant Bellvert ◽  
René Bally ◽  
Daniel Muller ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Agrobacterium Tumefaciens ◽  
Nitrite Reductase ◽  
Azospirillum Brasilense ◽  
High Performance ◽  
Positive Control ◽  
Shoot Biomass ◽  
E Coli ◽  
K 12 ◽  
The Impact

Plant-beneficial effects of bacteria are often underestimated, especially for well-studied strains associated with pathogenicity or originating from other environments. We assessed the impact of seed inoculation with the emblematic bacterial models Agrobacterium tumefaciens C58 (plasmid-cured) or Escherichia coli K-12 on maize seedlings in nonsterile soil. Compared with the noninoculated control, root biomass (with A. tumefaciens or E. coli) and shoot biomass (with A. tumefaciens) were enhanced at 10 days for ‘PR37Y15’ but not ‘DK315’, as found with the phytostimulator Azospirillum brasilense UAP-154 (positive control). In roots as well as in shoots, Agrobacterium tumefaciens and E. coli triggered similar (in PR37Y15) or different (in DK315) changes in the high-performance liquid chromatography profiles of secondary metabolites (especially benzoxazinoids), distinct from those of Azospirillum brasilense UAP-154. Genome sequence analysis revealed homologs of nitrite reductase genes nirK and nirBD and siderophore synthesis genes for Agrobacterium tumefaciens, as well as homologs of nitrite reductase genes nirBD and phosphatase genes phoA and appA in E. coli, whose contribution to phytostimulation will require experimental assessment. In conclusion, the two emblematic bacterial models had a systemic impact on maize secondary metabolism and resulted in unexpected phytostimulation of seedlings in the Azospirillum sp.-responsive cultivar.

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