A Consecutive Genome Engineering Method Reveals a New Phenotype and Regulation of Glucose and Glycerol Utilization in .

is a microorganism for production of 1,3-propanediol (1,3-PDO) and butanol, but suffers from lacking genetic tools for metabolic engineering to improve product titers. Furthermore, previous studies of have mainly focused on single genomic modification. The aim of this work is the development and application of a method for modification of multiple gene targets in the genome of .

Design of cell expansion processes for adherent-growing cells with mDoE-workflow.

Adherent cells, mammalian or human, are ubiquitous for production of viral vaccines, in gene therapy and in immuno-oncology. The development of a cell-expansion process with adherent cells is challenging as scale-up requires the expansion of the cell culture surface. Microcarrier (MC)-based cultures are still predominate.

Novel synthetic co-culture of and using CO and in situ generated H for the production of caproic acid via lactic acid.

is known to produce mainly acetate from CO and H, but the production of higher value chemicals is desired for the bioeconomy. Using chain-elongating bacteria, synthetic co-cultures have the potential to produce longer-chained products such as caproic acid. In this study, we present first results for a successful autotrophic co-cultivation of mutants and a wild-type strain in a stirred-tank bioreactor for the production of caproic acid from CO and H via the intermediate lactic acid.

Generation and evaluation of input values for computational analysis of transport processes within tissue cultures.

Techniques for tissue culture have seen significant advances during the last decades and novel 3D cell culture systems have become available. To control their high complexity, experimental techniques and their Digital Twins (modelling and computational tools) are combined to link different variables to process conditions and critical process parameters. This allows a rapid evaluation of the expected product quality.

Enhanced sequestration of carbon dioxide into calcium carbonate using pressure and a carbonic anhydrase from alkaliphilic .

CO in the atmosphere is a major contributor to global warming but at the same time it has the potential to be a carbon source for advanced biomanufacturing. To utilize CO, carbonic anhydrase has been identified as a key enzyme. Furthermore, attempts have been made to accelerate the sequestration via pressure.

A novel downstream process for highly pure 1,3-propanediol from an efficient fed-batch fermentation of raw glycerol by .

An efficient downstream process without prior desalination was developed for recovering 1,3-propanediol (1,3-PDO) with high purity and yield from broth of a highly productive fed-batch fermentation of raw glycerol by . After removal of biomass and proteins by ultrafiltration, and concentration by water evaporation, 1,3-PDO was directly recovered from the broth by vacuum distillation with continuous addition and regeneration of glycerol as a supporting agent. Inorganic salts in the fermentation broth were crystallized but well suspended by a continuous flow of glycerol during the distillation process, which prevented salt precipitation and decline of heat transfer.

Dynamic energy correlation analysis of aspartokinase III and alteration of allosteric regulation by manipulating energy transduction pathways.

Conformational change associated with allosteric regulation in a protein is ultimately driven by energy transformation. However, little is known about the latter process. In this work, we combined steered molecular dynamics simulations and sequence conservation analysis to investigate the conformational changes and energy transformation in the allosteric enzyme aspartokinase III (AK III) from .

Metabolomic and kinetic investigations on the electricity-aided production of butanol by strains.

In this contribution, we studied the effect of electro-fermentation on the butanol production of strains by a targeted metabolomics approach. Two strains were examined: an electrocompetent wild type strain (R525) and a mutant strain (dhaB mutant) lacking formation of 1,3-propanediol (PDO). The dhaB-negative strain was able to grow on glycerol without formation of PDO, but displayed a high initial intracellular NADH/NAD ratio which was lowered subsequently by upregulation of the butanol production pathway.

Regulation of pyruvate dehydrogenase complex related to lactate switch in CHO cells.

The metabolism of Chinese hamster ovary (CHO) cell lines is typically characterized by high rates of aerobic glycolysis with increased lactate formation, known as the "Warburg" effect. Although this metabolic state can switch to lactate consumption, the involved regulations of the central metabolism have only been partially studied so far. An important reaction transferring the lactate precursor, pyruvate, into the tricarboxylic acid cycle is the decarboxylation reaction catalyzed by the pyruvate dehydrogenase enzyme complex (PDC).

Improved electrocompetence and metabolic engineering of reveals a new regulation pattern of glycerol fermentation.

produces industrially valuable chemicals such as -butanol and 1,3-propanediol from fermentations of glycerol and glucose. Metabolic engineering for increased yields of selective compounds is not well established in this microorganism. In order to study carbon fluxes and to selectively increase butanol yields, we integrated the latest advances in genome editing to obtain an electrocompetent strain for further engineering.

A novel All-in-One electrolysis electrode and bioreactor enable better study of electrochemical effects and electricity-aided bioprocesses.

An autoclavable All-in-One electrolysis electrode in a rod shape assembly is developed as a new tool for bioelectrochemical systems and electricity-aided bioprocesses. It can replace the classic two-chamber bioelectrochemical system for electrolysis reactions, be inserted into conventional bioreactors and is easily adaptable as electrocatalytic surface or generator of super-fine bubbles (H and O) for bioconversion processes. Whereas the bioreactor itself functions as the working electrode chamber, a well-integrated inner counter electrode chamber enables water electrolysis without the normally encountered undesired ion-transfer effect.

Protein and pathway engineering for the biosynthesis of 5-hydroxytryptophan in .

The hydroxylation of tryptophan is an important reaction in the biosynthesis of natural products. 5-Hydroxytryptophan (5HTP) is not only an important compound for its pharmaceutical value but also because it is the precursor of other molecules, such as serotonin. In this study, we have extended the metabolism of an strain to produce 5HTP.

Analysis of intracellular metabolites of at high cell density with automated sampling and filtration and assessment of engineered enzymes for effective l-lysine production.

Engineering of enzymes and pathways is generally required for the development of efficient strains for bioproduction processes. To this end, quantitative and reliable data of intracellular metabolites are highly desired, but often not available, especially for conditions more close to industrial applications, i.e.

Weak cell cycle dependency but strong distortive effects of transfection with Lipofectamine 2000 in near-physiologically synchronized cell culture.

Previously, we reported a method to generate and validate cell cycle-synchronized cultures of multiple mammalian suspension cell lines under near-physiological conditions. This method was applied to elucidate the putative interdependencies of the cell cycle and recombinant protein expression in the human producer cell line HEK293s using Lipofectamine 2000 and the reporter plasmid pcDNA3.3 enhanced green fluorescent protein, destabilized using PEST sequence.

Glucose-mediated regulation of glycerol uptake in : Insights through transcriptomic analysis on dual substrate fermentation.

is a robust oleaginous yeast that can accumulate lipids to more than 70% of its dry cell mass. Even though it is extensively studied for its fermentation of substrates like glucose and glycerol, limited information is available about its metabolism of mixture of glucose and glycerol. During growth on mixture of glucose and glycerol a typical diauxic growth and higher lipid yields were observed.