Zymomonas mobilis genes for pyruvate decarboxylase (pdc) and alcohol dehydrogenase II (adhB) were integrated into the Escherichia coli chromosome within or near formate-lyase gene (pfl). Integration improved stability of Z. in E. coli, but further selection was required to increase expression. Spontaneous mutants selected resistance high level chloramphenicol that also expressed levels genes. Analogous increased expression on aldehyde indicator plates. These functionally equivalent previous...

Abstract Derivatives of Escherichia coli C were engineered to produce primarily succinate or malate in mineral salts media using simple fermentations (anaerobic stirred batch with pH control) without the addition plasmids foreign genes. This was done by a combination gene deletions (genetic engineering) and metabolic evolution over 2,000 generations growth‐based selection. After deletion central anaerobic fermentation genes ( ldhA , adhE ackA ), pathway for production remained as primary...

Escherichia coli TC44, a derivative of W3110, was engineered for the production pyruvate from glucose by combining mutations to minimize ATP yield, cell growth, and CO 2 (Δ atpFH Δ adhE sucA ) with that eliminate acetate [ poxB :: FRT (FLP recognition target) ackA ] fermentation products focA-pflB frdBC ldhA ). In mineral salts medium containing as sole carbon source, strain TC44(Δ converted yield 0.75 g per (77.9% theoretical yield; 1.2 liters –1 ·h A maximum 749 mM produced excess glucose....

During metabolic evolution to improve succinate production in Escherichia coli strains, significant changes cellular metabolism were acquired that increased energy efficiency two respects. The energy-conserving phosphoenolpyruvate (PEP) carboxykinase ( pck ), which normally functions the reverse direction (gluconeogenesis; glucose repressed) during oxidative of organic acids, evolved become major carboxylation pathway for production. Both PCK enzyme activity and gene expression levels...

The resistance of polylactide to biodegradation and the physical properties this polymer can be controlled by adjusting ratio L-lactic acid D-lactic acid. Although largest demand is for L enantiomer, substantial amounts both enantiomers are required bioplastics. We constructed derivatives Escherichia coli W3110 (prototrophic) as new biocatalysts production These strains (SZ40, SZ58, SZ63) require only mineral salts nutrients lack all plasmids antibiotic genes used during construction....

Derivatives of Escherichia coli C were previously described for succinate production by combining the deletion genes that disrupt fermentation pathways alternative products (ldhA::FRT, adhE::FRT, ackA::FRT, focA-pflB::FRT, mgsA, poxB) with growth-based selection increased ATP production. The resulting strain, KJ073, produced 1.2 mol per glucose in mineral salts medium acetate, malate, and pyruvate as significant co-products. KJ073 has been further improved removing residual recombinase sites...

Mutations causing constitutive synthesis of glutamine synthetase (GlnC − phenotype) were transferred from Klebsiella aerogenes into pneumoniae by P1-mediated transduction. Such GlnC strains K. have levels synthetase. Two three studied, each containing independently isolated mutations that confer the phenotype, continue to synthesize nitrogenase in presence NH 4 + . One strain, KP5069, produces 30% as much when grown 15 mM its absence. The phenotype allows under conditions completely repress...

Microbial processes for commodity chemicals have focused on reduced products and anaerobic conditions where substrate loss to cell mass CO 2 are minimal product yields high. To facilitate expansion into more oxidized chemicals, Escherichia coli W3110 was genetically engineered acetate production by using an approach that combines attributes of fermentative oxidative metabolism (rapid growth, external electron acceptor) a single biocatalyst. The resulting strain (TC36) converted 333 mM...

ABSTRACT Biomass-derived sugars, such as glucose, xylose, and other minor can be readily fermented to fuel ethanol commodity chemicals by the appropriate microbes. Due differences in optimum conditions for activity of fungal cellulases that are required depolymerization cellulose fermentable sugars growth fermentation characteristics current industrial microbes, simultaneous saccharification (SSF) is envisioned at not optimal cellulase activity, leading a higher-than-required cost SSF. We...

The efficient diversion of pyruvate from normal fermentative pathways to ethanol production in Klebsiella oxytoca M5A1 requires the expression Zymomonas mobilis genes encoding both decarboxylase and alcohol dehydrogenase. Final concentrations obtained with best recombinant, strain (pLOI555), were excess 40 g/liter an efficiency 0.48 g (xylose) 0.50 (glucose) per sugar, as compared a theoretical maximum 0.51 sugar. maximal volumetric productivity hour for sugars was 2.0 g/liter. This xylose...

Pretreatments such as dilute acid at elevated temperature are effective for the hydrolysis of pentose polymers in hemicellulose and also increase access enzymes to cellulose fibers. However, fermentation resulting syrups is hindered by minor reaction products furfural from dehydration. To mitigate this problem, four genetic traits have been identified that tolerance ethanol-producing Escherichia coli LY180 (strain W derivative): increased expression fucO , ucpA or pntAB deletion yqhD ....

ABSTRACT Low concentrations of furfural are formed as a side product during the dilute acid hydrolysis hemicellulose. Growth is inhibited by exposure to but resumes after complete reduction less toxic furfuryl alcohol. Growth-based selection was used isolate furfural-resistant mutant ethanologenic Escherichia coli LY180, designated strain EMFR9. Based on mRNA expression levels in parent and response challenge, genes encoding 12 oxidoreductases were found vary more than twofold (eight higher...

ABSTRACT Escherichia coli strains (KJ060 and KJ073) that were previously developed for succinate production have now been modified malate production. Many unexpected changes observed during this investigation. The initial strategy of deleting fumarase isoenzymes was ineffective, continued to accumulate. Surprisingly, a mutation in fumarate reductase alone sufficient redirect carbon flow into even the presence fumarase. Further deletions needed inactivate malic enzymes (typically...

A wide variety of commercial products can be potentially made from monomeric sugars produced by the dilute acid hydrolysis lignocellulosic biomass. However, this process is accompanied side such as furfural that hinder microbial growth and fermentation. To investigate mechanism inhibition, mRNA microarrays an ethanologenic strain Escherichia coli (LY180) were compared immediately prior to 15 min after a moderate challenge. Expression genes regulators associated with biosynthesis cysteine...

The genes encoding the two structural subunits of Escherichia coli hydrogenase 2 (HYD2) have been cloned and sequenced. They occur in an operon (hyb) which contains seven open reading frames. An hyb deletion mutant (strain AP3) failed to grown on dihydrogen-fumarate medium also produced very low levels HYD1. All frames are required for restoration wild-type active HYD2 AP3. was mapped at 65 min E. chromosome.

Deletion mutants of Escherichia coli specific for hydrogenase isoenzyme 1 (HYD1) have been constructed and characterized. The hya operon, which contains genes the two HYD1 structural subunits four additional genes, was mapped at 22 min on E. chromosome. total activities HYD1-negative mutant wild-type strains were similar. However, formate dehydrogenase activity associated with hydrogen lyase pathway lower in mutant. (strain AP1), complemented only (hyaAB), produced antigenically identifiable...

During anaerobic growth of bacteria, organic intermediates metabolism, such as pyruvate or its derivatives, serve electron acceptors to maintain the overall redox balance. Under these conditions, ATP needed for cell is derived from substrate-level phosphorylation. In Escherichia coli, conversion glucose yields 2 net ATPs, while metabolism a pentose, xylose, only 0.67 per xylose due need one (each) transport and xylulose fermentative growth, E. coli produces equimolar amounts acetate ethanol...

Conversion of lignocellulosic feedstocks to ethanol requires microorganisms that effectively ferment both hexose and pentose sugars. Towards this goal, recombinant organisms have been developed in which heterologous genes were added platform such as Saccharomyces cerevisiae, Zymomonas mobilis, Escherichia coli. Using a novel approach relies only on native enzymes, we homoethanologenic alternative, coli strain SE2378. This mutant ferments glucose or xylose with yield 82% under anaerobic...

Abstract Escherichia coli KO11 (parent) and LY01 (mutant) have been engineered for the production of ethanol. Gene arrays were used to identify expression changes that occurred in mutant, LY01, during directed evolution improve ethanol tolerance (defined as extent growth presence added ethanol). Expression levels 205 (5%) ORFs found differ significantly ( p < 0.10) between under each six different conditions 0.000001). Statistical evaluation differentially expressed genes according...