Poly(3-hydroxybutyrate) synthesis from glycerol by a recombinant Escherichia coli arcA mutant in fed-batch microaerobic cultures (Applied Microbiology and Biotechnology DOI: 10.1007/s00253-007-1255-7)

Fil:Pettinari, M.J. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina.Fil:Galvagno, M.A. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina.Fil:Méndez, B.S. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina

Poly(3-hydroxybutyrate) synthesis by recombinant Escherichia coli arcA mutants in microaerobiosis

We assessed the effects of different arcA mutations on poly(3- hydroxybutyrate) (PHB) synthesis in recombinant Escherichia coli strains carrying the pha synthesis genes from Azotobacter sp. strain FA8. The arcA mutations used were an internal deletion and the arcA2 allele, a leaky mutation for some of the characteristics of the Arc phenotype which confers high respiratory capacity. PHB synthesis was not detected in the wild-type strain in shaken flask cultures under low-oxygen conditions, while ArcA mutants gave rise to polymer accumulation of up to 24% of their cell dry weight. When grown under microaerobic conditions in a bioreactor, the arcA deletion mutant reached a PHB content of 27% ± 2%. Under the same conditions, higher biomass and PHB concentrations were observed for the strain bearing the arcA2 allele, resulting in a PHB content of 35% ± 3%. This strain grew in a simple medium at a specific growth rate of 0.69 ± 0.07 h-1, whereas the deletion mutant needed several nutritional additives and snowed a specific growth rate of 0.56 ± 0.06 h-1. The results presented here suggest that arcA mutations could play a role in heterologous PHB synthesis in microaerobiosis. Copyright © 2006, American Society for Microbiology. All Rights Reserved.Fil:Pettinari, M.J. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina.Fil:Galvagno, M.A. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina.Fil:Méndez, B.S. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina

Ethanol synthesis from glycerol by Escherichia coli redox mutants expressing adhE from Leuconostoc mesenteroides

Aims: Analysis of the physiology and metabolism of Escherichia coli arcA and creC mutants expressing a bifunctional alcohol-acetaldehyde dehydrogenase from Leuconostoc mesenteroides growing on glycerol under oxygen-restricted conditions. The effect of an ldhA mutation and different growth medium modifications was also assessed. Methods and Results: Expression of adhE in E. coli CT1061 [arcA creC(Con)] resulted in a 1·4-fold enhancement in ethanol synthesis. Significant amounts of lactate were produced during micro-oxic cultures and strain CT1061LE, in which fermentative lactate dehydrogenase was deleted, produced up to 6·5 ± 0·3 g l-1 ethanol in 48 h. Escherichia coli CT1061LE derivatives resistant to >25 g l-1 ethanol were obtained by metabolic evolution. Pyruvate and acetaldehyde addition significantly increased both biomass and ethanol concentrations, probably by overcoming acetyl-coenzyme A (CoA) shortage. Yeast extract also promoted growth and ethanol synthesis, and this positive effect was mainly attributable to its vitamin content. Two-stage bioreactor cultures were conducted in a minimal medium containing 100 μg l-1 calcium d-pantothenate to evaluate oxic acetyl-CoA synthesis followed by a switch into fermentative conditions. Ethanol reached 15·4 ± 0·9 g l-1 with a volumetric productivity of 0·34 ± 0·02 g l-1 h-1. Conclusions: Escherichia coli responded to adhE over-expression by funnelling carbon and reducing equivalents into a highly reduced metabolite, ethanol. Acetyl-CoA played a key role in micro-oxic ethanol synthesis and growth. Significance and Impact of the Study: Insight into the micro-oxic metabolism of E. coli growing on glycerol is essential for the development of efficient industrial processes for reduced biochemicals production from this substrate, with special relevance to biofuels synthesis. © 2010 The Society for Applied Microbiology.Fil:Pettinari, M.J. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina.Fil:Méndez, B.S. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina.Fil:Galvagno, M.A. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina

New recombinant Escherichia coli strain tailored for the production of poly(3-hydroxybutyrate) from agroindustrial by-products

A recombinant E. coli strain (K24K) was constructed and evaluated For poly(3-hydroxybutyrate) (PHB) production from whey and corn steep liquor as main carbon and nitrogen sources. This strain bears the pha biosynthetic genes from Azotobacter sp. strain FA8 expressed from a T5 promoter under the control of the lactose operator. K24K does not produce the lactose repressor, ensuring constitutive expression of genes involved in lactose transport and utilization. PHB was efficiently produced by the recombinant strain grown aerobically in fed-batch cultures in a laboratory scale bioreactor on a semisynthetic medium supplemented with the agroindustrial by-products. After 24 h, cells accumulated PHB to 72.9% of their cell dry weight, reaching a volumetric productivity of 2.13 g PHB per liter per hour. Physical analysis of PHB recovered from the recombinants showed that its molecular weight was similar to that of PHB produced by Azotobacter sp. strain FA8 and higher than that of the polymer from Cupriavidus necator and that its glass transition temperature was approximately 20°C higher than those of PHBs from the natural producer strains. Copyright © 2006, American Society for Microbiology. All Rights Reserved.Fil:De Almeida, A. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina.Fil:Galvagno, M.A. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina.Fil:Pettinari, M.J. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina

Extracorporeal life support in mitral papillary muscle rupture: Outcome of multicenter study

Background: Post-acute myocardial infarction papillary muscle rupture (post-AMI PMR) may present variable clinical scenarios and degree of emergency due to result of cardiogenic shock. Veno-arterial extracorporeal life support (V-A ECLS) has been proposed to improve extremely poor pre- or postoperative conditions. Information in this respect is scarce.Methods: From the CAUTION (meChanical complicAtion of acUte myocardial infarcTion: an InternatiOnal multiceNter cohort study) database (16 different Centers, data from 2001 to 2018), we extracted adult patients who were surgically treated for post-AMI PMR and underwent pre- or/and postoperative V-A ECLS support. The end-points of this study were in-hospital survival and ECLS complications.Results: From a total of 214 post-AMI PMR patients submitted to surgery, V-A ECLS was instituted in 23 (11%) patients. The median age was 61.7 years (range 46-81 years). Preoperatively, ECLS was commenced in 10 patients (43.5%), whereas intra/postoperative in the remaining 13. The most common V-A ECLS indication was post-cardiotomy shock, followed by preoperative cardiogenic shock and cardiac arrest. The median duration of V-A ECLS was 4 days. V-A ECLS complications occurred in more than half of the patients. Overall, in-hospital mortality was 39.2% (9/23), compared to 22% (42/219) for the non-ECLS group.Conclusions: In post-AMI PMR patients, V-A ECLS was used in almost 10% of the patients either to promote bridge to surgery or as postoperative support. Further investigations are required to better evaluate a potential for increased use and its effects of V-A ECLS in such a context based on the still high perioperative mortality