Elucidating regulation of polyhydroxyalkanoate metabolism in Ralstonia eutropha: Identification of transcriptional regulators from phasin and depolymerase genes.

Despite the ever-growing research interest in polyhydroxyalkanoates (PHAs) as green plastic alternatives, our understanding of the regulatory mechanisms governing PHA synthesis, storage, and degradation in the model organism Ralstonia eutropha remains limited. Given its importance for central carbon metabolism, PHA homeostasis is probably controlled by a complex network of transcriptional regulators. Understanding this fine-tuning is the key for developing improved PHA production strains thereby boosting the application of PHAs.

Recent insights into the world of dual-function bacterial sRNAs.

Dual-function sRNAs refer to a small subgroup of small regulatory RNAs that merges base-pairing properties of antisense RNAs with peptide-encoding properties of mRNA. Both functions can be part of either same or in another metabolic pathway. Here, we want to update the knowledge of to the already known dual-function sRNAs and review the six new sRNAs found since 2017 regarding their structure, functional mechanisms, evolutionary conservation, and role in the regulation of distinct biological/physiological processes.

Improved preculture management for Cupriavidus necator cultivations.

Objectives: Research on hydrogenases from Cupriavidus necator has been ongoing for more than two decades and still today the common methods for culture inoculation are used. These methods were never adapted to the requirements of modified bacterial strains, resulting in different physiological states of the bacteria in the precultures, which in turn lead prolonged and different lag-phases.

Results: In order to obtain uniform and always equally fit precultures for inoculation, we have established in this study an optimized protocol for precultures of the derivative of C.

Implementation of a high cell density fed-batch for heterologous production of active [NiFe]-hydrogenase in Escherichia coli bioreactor cultivations.

Background: O-tolerant [NiFe]-hydrogenases offer tremendous potential for applications in H-based technology. As these metalloenzymes undergo a complicated maturation process that requires a dedicated set of multiple accessory proteins, their heterologous production is challenging, thus hindering their fundamental understanding and the development of related applications. Taking these challenges into account, we selected the comparably simple regulatory [NiFe]-hydrogenase (RH) from Cupriavidus necator as a model for the development of bioprocesses for heterologous [NiFe]-hydrogenase production.

High-Yield Production of Catalytically Active Regulatory [NiFe]-Hydrogenase From in .

Hydrogenases are biotechnologically relevant metalloenzymes that catalyze the reversible conversion of molecular hydrogen into protons and electrons. The O-tolerant [NiFe]-hydrogenases from (formerly ) are of particular interest as they maintain catalysis even in the presence of molecular oxygen. However, to meet the demands of biotechnological applications and scientific research, a heterologous production strategy is required to overcome the low production yields in their native host.

Production of soluble regulatory hydrogenase from Ralstonia eutropha in Escherichia coli using a fed-batch-based autoinduction system.

Background: Autoinduction systems can regulate protein production in Escherichia coli without the need to monitor cell growth or add inducer at the proper time following culture growth. Compared to classical IPTG induction, autoinduction provides a simple and fast way to obtain high protein yields. In the present study, we report on the optimization process for the enhanced heterologous production of the Ralstonia eutropha regulatory hydrogenase (RH) in E.

Optimization of Culture Conditions for Oxygen-Tolerant Regulatory [NiFe]-Hydrogenase Production from H16 in .

Hydrogenases are abundant metalloenzymes that catalyze the reversible conversion of molecular H into protons and electrons. Important achievements have been made over the past two decades in the understanding of these highly complex enzymes. However, most hydrogenases have low production yields requiring many efforts and high costs for cultivation limiting their investigation.

Heterologous Hydrogenase Overproduction Systems for Biotechnology-An Overview.

Hydrogenases are complex metalloenzymes, showing tremendous potential as H-converting redox catalysts for application in light-driven H production, enzymatic fuel cells and H-driven cofactor regeneration. They catalyze the reversible oxidation of hydrogen into protons and electrons. The apo-enzymes are not active unless they are modified by a complicated post-translational maturation process that is responsible for the assembly and incorporation of the complex metal center.

A UV/Vis Spectroscopy-Based Assay for Monitoring of Transformations Between Nucleosides and Nucleobases.

Efficient reaction monitoring is crucial for data acquisition in kinetic and mechanistic studies. However, for conversions of nucleosides to their corresponding nucleobases, as observed in enzymatically catalyzed nucleoside phosphorylation reactions, the current analytical arsenal does not meet modern requirements regarding cost, speed of analysis and high throughput. Herein, we present a UV/Vis spectroscopy-based assay employing an algorithm for spectral unmixing in a 96-well plate format.

Bioactive Secondary Metabolites from : A Comprehensive Review.

is widely underappreciated for its inherent biosynthetic potential. This report comprehensively summarizes the known bioactive secondary metabolites from and highlights potential applications as plant pathogen control agents, drugs, and biosurfactants. is well known for the production of cyclic lipopeptides exhibiting strong surfactant and antimicrobial activities, such as surfactins, iturins, and fengycins.

A new role for CsrA: promotion of complex formation between an sRNA and its mRNA target in .

CsrA is a widely conserved, abundant small RNA binding protein that has been found in and other Gram-negative bacteria where it is involved in the regulation of carbon metabolism, biofilm formation and virulence. CsrA binds to single-stranded GGA motifs around the SD sequence of target mRNAs where it inhibits or activates translation or influences RNA processing. Small RNAs like CsrB or CsrC containing 13-22 GGA motifs can sequester CsrA, thereby abrogating the effect of CsrA on its target mRNAs.

Antibacterial and anticancer activities of orphan biosynthetic gene clusters from Atlantis II Red Sea brine pool.

Background: Cancer and infectious diseases are problematic because of continuous emergence of drug resistance. One way to address this enormous global health threat is bioprospecting the unlikeliest environments, such as extreme marine niches, which have tremendous biodiversity that is barely explored. One such environment is the Red Sea brine pool, Atlantis II Deep (ATII).

Automated Cell Treatment for Competence and Transformation of in a High-Throughput Quasi-Turbidostat Using Microtiter Plates.

Metabolic engineering and genome editing strategies often lead to large strain libraries of a bacterial host. Nevertheless, the generation of competent cells is the basis for transformation and subsequent screening of these strains. While preparation of competent cells is a standard procedure in flask cultivations, parallelization becomes a challenging task when working with larger libraries and liquid handling stations as transformation efficiency depends on a distinct physiological state of the cells.

Characterization of the interaction between the small RNA-encoded peptide SR1P and GapA from Bacillus subtilis.

Small regulatory RNAs (sRNAs) are the most prominent post-transcriptional regulators in all kingdoms of life. A few of them, e.g.

Dual-function small regulatory RNAs in bacteria.

Dual-function sRNAs are a subgroup of small regulatory RNAs that act on the one hand as base-pairing sRNAs to inhibit or activate target gene expression and on the other hand as peptide-encoding mRNAs that function either in the same or in another metabolic pathway. Here, we review and compare the five currently known and intensively characterized dual-function sRNAs with regard to their two functions, their biological role, their evolutionary conservation and their requirements for RNA chaperones. Furthermore, we summarize the data available on five potential dual-function sRNAs, whose base-pairing function is well established whereas the role of their encoded peptides has not yet been elucidated.

Dual-function sRNA encoded peptide SR1P modulates moonlighting activity of B. subtilis GapA.

SR1 is a dual-function sRNA from B. subtilis that acts as a base-pairing regulatory RNA and as a peptide-encoding mRNA. Both functions of SR1 are highly conserved.

SR1--a small RNA with two remarkably conserved functions.

SR1 is a dual-function sRNA that acts as a base-pairing regulatory RNA on the ahrC mRNA and as a peptide-encoding mRNA on the gapA operon. The SR1-encoded peptide SR1P binds GapA thereby stabilizing gapA mRNA. Under glycolytic conditions, SR1 transcription is repressed by CcpN and CcpA.

Construction of a modular plasmid family for chromosomal integration in Bacillus subtilis.

The investigation of molecular processes involves the generation of knockout strains, the determination of promoter strength and protein overexpression. Here, we report the construction of the multifunctional pMG expression vector family for integration into the Bacillus subtilis chromosome that allows gene expression under single copy conditions. The pMG family enables a rapid exchange of all features for integration, selection and gene expression with or without N-terminal strep-tags.

A dual-function sRNA from B. subtilis: SR1 acts as a peptide encoding mRNA on the gapA operon.

Small non-coding RNAs (sRNAs) have been found to regulate gene expression in all three kingdoms of life. So far, relatively little is known about sRNAs from Gram-positive bacteria. SR1 is a regulatory sRNA from the Bacillus subtilis chromosome that inhibits by base-pairing translation initiation of ahrC mRNA encoding a transcriptional activator of the arginine catabolic operons.