Targeted Cell Labeling and Sorting of Prokaryotes for Cultivation and Omics Approaches.

To date, the vast majority of prokaryotic organisms escapes detailed characterization because they cannot be isolated in axenic cultures. These organisms are referred to as microbial dark matter. Targeted labelling and sorting of these microorganisms pave the way for single-cell, enrichment, or cultivation approaches. In this review, we describe an array of different methods ranging from labeling-free to specific labelling techniques. In addition, different cell sorting methods and their combinations with targeting strategies are summarized and downstream applications like sequencing and cultivation are reviewed. Recent advances, challenges, and limitations of the particular methods are discussed with respect to cell viability, genome integrity as well as throughput, in order to help researchers select the most suitable methods for their specific research questions.

Bioaugmentation with marine sediment-derived microbial consortia in mesophilic anaerobic digestion for enhancing methane production under ammonium or salinity stress.

Ammonium (NH4+) and salinity (NaCl) inhibit CH4 production in anaerobic digestion. However, whether bioaugmentation using marine sediment-derived microbial consortia can relieve the inhibitory effects of NH4+ and NaCl stresses on CH4 production remains unclear. Thus, this study evaluated the effectiveness of bioaugmentation using marine sediment-derived microbial consortia in alleviating the inhibition of CH4 production under NH4+ or NaCl stress and elucidated the underlying mechanisms. Batch anaerobic digestion experiments under 5 gNH4-N/L or 30 g/L NaCl were performed with or without augmentation using two marine sediment-derived microbial consortia pre-acclimated to high NH4+ and NaCl. Compared with non-bioaugmentation, bioaugmentation reinforced CH4 production. Network analysis revealed the joint effects of microbial connections by Methanoculleus, which promoted the efficient consumption of propionate accumulated under NH4+ and NaCl stresses. In conclusion, bioaugmentation with pre-acclimated marine sediment-derived microbial consortia can mitigate the inhibition under NH4+ or NaCl stress and enhance CH4 production in anaerobic digestion.

Psychrophile-based simple biocatalysts for effective coproduction of 3-hydroxypropionic acid and 1,3-propanediol.

3-Hydroxypropionic acid (3-HP) and 1,3-propanediol (1,3-PDO) have tremendous potential markets in many industries. This study evaluated the simultaneous biosynthesis of the 2 compounds using the new psychrophile-based simple biocatalyst (PSCat) reaction system. The PSCat method is based on the expression of glycerol dehydratase, 1,3-propanediol dehydrogenase, and aldehyde dehydrogenase from Klebsiella pneumoniae in Shewanella livingstonensis Ac10 and Shewanella frigidimarina DSM 12253, individually. Heat treatment at 45 °C for 15 min deactivated the intracellular metabolic flux, and the production process was started after adding substrate, cofactor, and coenzyme. In the solo production process after 1 h, the maximum production of 3-HP was 62.0 m m. For 1,3-PDO, the maximum production was 25.0 m m. In the simultaneous production process, productivity was boosted, and the production of 3-HP and 1,3-PDO increased by 13.5 and 4.9 m m, respectively. Hence, the feasibility of the individual production and the simultaneous biosynthesis system were verified in the new PSCat approach.

Efficient production of 1,3-propanediol by psychrophile-based simple biocatalysts in Shewanella livingstonensis Ac10 and Shewanella frigidimarina DSM 12253.

1,3-Propanediol (1,3-PDO) is a valuable compound with a large potential market in many industries. This study aimed to evaluate the abilities of the Psychrophile-based Simple bioCatalyst (PSCat) reaction system to biosynthesize 1,3-PDO. This biocatalyst has a potential platform that replaces the chemical-based production counterparts. The two genes involved in the metabolic pathway were expressed both individually and together in the psychrophilic host bacterium. The intracellular metabolic flux was deactivated using heat treatment, at 45 °C for 15 min. After individual gene expression (25.0 mM), 1,3-PDO productivity of the cells increased by approximately 2.5 times, in comparison to when genes were expressed together (10.2 mM). Productivity was boosted (31.1 mM) when the cofactor regeneration system was activated in the biocatalyst. Hence, both the ability of individual gene expression and the cofactor regeneration system were verified in the PSCat approach. Nonetheless, further research is necessary to develop and optimize this process for industrial production.

The effect of structure on improvement of the PNA Young modulus: A study of steered molecular dynamics.

Prefoldin is a molecular chaperone and acts as a nano-actuator in cargo carriage and drug delivery for disease treatment. Investigating the mechanical properties of nano-actuator helps predict its behavior and measure its performance under various environmental conditions, like external forces that are applied. Accordingly, this paper investigates the elastic properties of the Prefoldin nano-actuator (PNA), specifically its Young modulus and the structural changes on a microscopic scale. For this purpose, three structurally different PNAs obtained from Protein Data Bank (PDB) and previous studies of our research team have been used. The selected three-tentacles Prefoldin are analyzed via the series of steered molecular dynamics simulations (SMD) based on the theory of Two Springs in Series. The simulation is applied in the velocity of 0.1, 0.05, and 0.01. Due to differences in the structure of the Prefoldin, PNAs exhibited different behaviours at various pull rates. Also, the analysis showed different values of Young's modulus for the PNA tentacles in the interval of (2.5-4 GPa). Understanding the mechanical properties of a Prefoldin nano actuator allows for a closer examination of its application in transportation the pathogenic cargos and intelligent drug delivery.

Force Sensing in Nanoscale: Integration of Virtual Spring for Sensing the Interactions of ß-Amyloid Grabbed by Prefoldin.

Analysis of atomic forces in molecular scale including the reciprocal internal force directions, quantity, and Young's modulus, and more detailed analysis via diagrams, is one of the most current research topics. This research aims to analyze the bilateral forces applied between a nano-actuator and a cargo from a new perspective. While virtual springs have been used as a tool to convey forces in the previous steered molecular dynamics simulations, this study uses the spring as a sensor for measuring the internal forces. This study introduces atomic sensing via the steered molecular dynamics method. Following the previous studies, the protein employed in this study is the mutated Archetype Prefoldin being used to control the pathogenic cargo beta-amyloid (Alzheimer's). The powerful Gromacs software carries out the simulations for the calculation of the total force and force for each branch. The simulation results illustrate the total force between the cargo and nano-actuator is ~ 2.8 nN, while each branch needs a force of ~ 1.2-1.5 nN to release the cargo. The results demonstrate the validity of the method and applicability of the virtual sensor for assessing the microscopic forces. This investigation is a pioneer study for the advent of the sensor as an assessment tool for the mechanical analysis and precise atomic force studies.