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 .

Shewanella oneidensis: Biotechnological Application of Metal-Reducing Bacteria.

What is an unconventional organism in biotechnology? The γ-proteobacterium Shewanella oneidensis might fall into this category as it was initially established as a laboratory model organism for a process that was not seen as potentially interesting for biotechnology. The reduction of solid-state extracellular electron acceptors such as iron and manganese oxides is highly relevant for many biogeochemical cycles, although it turned out in recent years to be quite relevant for many potential biotechnological applications as well. Applications started with the production of nanoparticles and dramatically increased after understanding that electrodes in bioelectrochemical systems can also be used by these organisms.

Comparative analysis of the influence of BpfA and BpfG on biofilm development and current density in Shewanella oneidensis under oxic, fumarate- and anode-respiring conditions.

Biofilm formation by Shewanella oneidensis has been extensively studied under oxic conditions; however, relatively little is known about biofilm formation under anoxic conditions and how biofilm architecture and composition can positively influence current generation in bioelectrochemical systems. In this study, we utilized a recently developed microfluidic biofilm analysis setup with automated 3D imaging to investigate the effects of extracellular electron acceptors and synthetic modifications to the extracellular polymeric matrix on biofilm formation. Our results with the wild type strain demonstrate robust biofilm formation even under anoxic conditions when fumarate is used as the electron acceptor.

Elucidating the development of cooperative anode-biofilm-structures.

Microbial electrochemical systems are a highly versatile platform technology with a particular focus on the interplay of chemical and electrical energy conversion and offer immense potential for a sustainable bioeconomy. The industrial realization of this potential requires a critical focus on biofilm optimization if performance is to be controlled over a long period of time. Moreover, the aspect and influence of cooperativity has to be addressed as many applied anodic bioelectrochemical systems will most likely be operated with a diversity of interacting microbial species.

When Cortical Bone Matrix Properties Are Indiscernible between Elderly Men with and without Type 2 Diabetes, Fracture Resistance Follows Suit.

Type 2 diabetes mellitus (T2DM) is a metabolic disease affecting bone tissue and leading to increased fracture risk in men and women, independent of bone mineral density (BMD). Thus, bone material quality (i.e.

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.

Microbe-Anode Interactions: Comparing the impact of genetic and material engineering approaches to improve the performance of microbial electrochemical systems (MES).

Microbial electrochemical systems (MESs) are a highly versatile platform technology with a particular focus on power or energy production. Often, they are used in combination with substrate conversion (e.g.

Scaling-down biopharmaceutical production processes via a single multi-compartment bioreactor (SMCB).

Biopharmaceutical production processes often use mammalian cells in bioreactors larger than 10,000 L, where gradients of shear stress, substrate, dissolved oxygen and carbon dioxide, and pH are likely to occur. As former tissue cells, producer cell lines such as Chinese hamster ovary (CHO) cells sensitively respond to these mixing heterogeneities, resulting in related scenarios being mimicked in scale-down reactors. However, commonly applied multi-compartment approaches comprising multiple reactors impose a biasing shear stress caused by pumping.

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.

Sprayable biofilm - Agarose hydrogels as 3D matrix for enhanced productivity in bioelectrochemical systems.

Bio-based energy production utilizing renewable resources can be realized by exoelectrogenic organisms and their application in bioelectrochemical systems (BES). These organisms catalyze the direct conversion of chemical into electrical energy and are already widely used in bioelectronics and biosensing. However, the biofilm-electrode interaction is a factor that limits sufficient space-time-yields for industrial applications.

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.

Improvement of glycine biosynthesis from one-carbon compounds and ammonia catalyzed by the glycine cleavage system in vitro.

Glycine cleavage system (GCS) plays a central role in one-carbon (C1) metabolism and receives increasing interest as a core part of the recently proposed reductive glycine pathway (rGlyP) for assimilation of CO and formate. Despite decades of research, GCS has not yet been well understood and kinetic data are barely available. This is to a large degree because of the complexity of GCS, which is composed of four proteins (H, T, P, and L) and catalyzes reactions involving different substrates and cofactors.

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).

Microbubble enhanced mass transfer efficiency of CO capture utilizing aqueous triethanolamine for enzymatic resorcinol carboxylation.

The present study focuses on the aeration of aqueous triethanolamine acting as reaction medium for biocatalytic carboxylations. For enhancing mass transfer in a bubble column reactor, microbubble aeration is applied and compared to conventional macrobubble aeration. Application of a 0.

A multi-omic screening approach for the discovery of thermoactive glycoside hydrolases.

Next-generation sequencing and computational biology have facilitated the implementation of new combinatorial screening approaches to discover novel enzymes of biotechnological interest. In this study, we describe the successful establishment of a multi-omic approach for the identification of thermostable hydrolase-encoding genes by determination of gene expression levels. We applied this combinatorial approach using an anaerobic enrichment culture from an Azorean hot spring sample grown on green coffee beans as recalcitrant substrate.

Biomass-degrading glycoside hydrolases of archaeal origin.

During the last decades, the impact of hyperthermophiles and their enzymes has been intensively investigated for implementation in various high-temperature biotechnological processes. Biocatalysts of hyperthermophiles have proven to show extremely high thermo-activities and thermo-stabilities and are identified as suitable candidates for numerous industrial processes with harsh conditions, including the process of an efficient plant biomass pretreatment and conversion. Already-characterized archaea-originated glycoside hydrolases (GHs) have shown highly impressive features and numerous enzyme characterizations indicated that these biocatalysts show maximum activities at a higher temperature range compared to bacterial ones.

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.

Formulation of organic and inorganic hydrogel matrices for immobilization of β-glucosidase in microfluidic platform.

The aim of this study was to formulate silica and alginate hydrogels for immobilization of β-glucosidase. For this purpose, enzyme kinetics in hydrogels were determined, activity of immobilized enzymes was compared with that of free enzyme, and structures of silica and alginate hydrogels were characterized in terms of surface area and pore size. The addition of polyethylene oxide improved the mechanical strength of the silica gels and 68% of the initial activity of the enzyme was preserved after immobilizing into tetraethyl orthosilicate-polyethylene oxide matrix where the relative activity in alginate beads was 87%.

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.