Activation of formate hydrogen-lyase via expression of uptake [NiFe]-hydrogenase in Escherichia coli BL21(DE3)

Background: Several recent studies have reported successful hydrogen (H-2) production achieved via recombinant expression of uptake [NiFe]-hydrogenases from Hydrogenovibrio marinus, Rhodobacter sphaeroides, and Escherichia coli (hydrogenase-1) in E. coli BL21(DE3), a strain that lacks H-2-evolving activity. However, there are some unclear points that do not support the conclusion that the recombinant hydrogenases are responsible for the in vivo H-2 production. Results: Unlike wild-type BL21(DE3), the recombinant BL21(DE3) strains possessed formate hydrogen-lyase (FHL) activities. Through experiments using fdhF (formate dehydrogenase-H) or hycE (hydrogenase-3) mutants, it was shown that H-2 production was almost exclusively dependent on FHL. Upon expression of hydrogenase, extracellular formate concentration was changed even in the mutant strains lacking FHL, indicating that formate metabolism other than FHL was also affected. The two subunits of H. marinus uptake [NiFe]-hydrogenase could activate FHL independently of each other, implying the presence of more than two different mechanisms for FHL activation in BL21(DE3). It was also revealed that the signal peptide in the small subunit was essential for activation of FHL via the small subunit. Conclusions: Herein, we demonstrated that the production of H-2 was indeed induced via native FHL activated by the expression of recombinant hydrogenases. The recombinant strains with [NiFe]-hydrogenase appear to be unsuitable for practical in vivo H-2 production due to their relatively low H-2 yields and productivities. We suggest that an improved H-2-producing cell factory could be designed by constructing a well characterized and overproduced synthetic H-2 pathway and fully activating the native FHL in BL21(DE3).1122Ysciescopu

Improved production of biohydrogen in light-powered Escherichia coli by co-expression of proteorhodopsin and heterologous hydrogenase

<p>Abstract</p> <p>Background</p> <p>Solar energy is the ultimate energy source on the Earth. The conversion of solar energy into fuels and energy sources can be an ideal solution to address energy problems. The recent discovery of proteorhodopsin in uncultured marine γ-proteobacteria has made it possible to construct recombinant <it>Escherichia coli </it>with the function of light-driven proton pumps. Protons that translocate across membranes by proteorhodopsin generate a proton motive force for ATP synthesis by ATPase. Excess protons can also be substrates for hydrogen (H₂) production by hydrogenase in the periplasmic space. In the present work, we investigated the effect of the co-expression of proteorhodopsin and hydrogenase on H₂production yield under light conditions.</p> <p>Results</p> <p>Recombinant <it>E. coli </it>BL21(DE3) co-expressing proteorhodopsin and [NiFe]-hydrogenase from <it>Hydrogenovibrio marinus </it>produced ~1.3-fold more H₂in the presence of exogenous retinal than in the absence of retinal under light conditions (70 μmole photon/(m²·s)). We also observed the synergistic effect of proteorhodopsin with endogenous retinal on H₂production (~1.3-fold more) with a dual plasmid system compared to the strain with a single plasmid for the sole expression of hydrogenase. The increase of light intensity from 70 to 130 μmole photon/(m²·s) led to an increase (~1.8-fold) in H₂production from 287.3 to 525.7 mL H₂/L-culture in the culture of recombinant <it>E. coli </it>co-expressing hydrogenase and proteorhodopsin in conjunction with endogenous retinal. The conversion efficiency of light energy to H₂achieved in this study was ~3.4%.</p> <p>Conclusion</p> <p>Here, we report for the first time the potential application of proteorhodopsin for the production of biohydrogen, a promising alternative fuel. We showed that H₂production was enhanced by the co-expression of proteorhodopsin and [NiFe]-hydrogenase in recombinant <it>E. coli </it>BL21(DE3) in a light intensity-dependent manner. These results demonstrate that <it>E. coli </it>can be applied as light-powered cell factories for biohydrogen production by introducing proteorhodopsin.</p

Implications of Personality on Cognitive Workload, Affect, and Task Performance in Remote Robot Control

This paper explores how the personality traits of robot operators can influence their task performance during remote control of robots. It is essential to explore the impact of personal dispositions on information processing, both directly and indirectly, when working with robots on specific tasks. To investigate this relationship, we utilize the open-access multi-modal dataset MOCAS to examine the robot operator's personality traits, affect, cognitive load, and task performance. Our objective is to confirm if personality traits have a total effect, including both direct and indirect effects, that could significantly impact the performance levels of operators. Specifically, we examine the relationship between personality traits such as extroversion, conscientiousness, and agreeableness, and task performance. We conduct a correlation analysis between cognitive load, self-ratings of workload and affect, and quantified individual personality traits along with their experimental scores. The findings show that personality traits do not have a total effect on task performance.Comment: 8 pages, 6 figures, accepted to IROS 2023. A link to a supplementary video is in the abstrac

Production of biohydrogen by recombinant expression of [NiFe]-hydrogenase 1 in Escherichia coli

<p>Abstract</p> <p>Background</p> <p>Hydrogenases catalyze reversible reaction between hydrogen (H₂) and proton. Inactivation of hydrogenase by exposure to oxygen is a critical limitation in biohydrogen production since strict anaerobic conditions are required. While [FeFe]-hydrogenases are irreversibly inactivated by oxygen, it was known that [NiFe]-hydrogenases are generally more tolerant to oxygen. The physiological function of [NiFe]-hydrogenase 1 is still ambiguous. We herein investigated the H₂production potential of [NiFe]-hydrogenase 1 of <it>Escherichia coli in vivo </it>and <it>in vitro</it>. The <it>hya</it>A and <it>hya</it>B genes corresponding to the small and large subunits of [NiFe]-hydrogenase 1 core enzyme, respectively, were expressed in BL21, an <it>E. coli </it>strain without H₂producing ability.</p> <p>Results</p> <p>Recombinant BL21 expressing [NiFe]-hydrogenase 1 actively produced H₂(12.5 mL H₂/(h·L) in 400 mL glucose minimal medium under micro-aerobic condition, whereas the wild type BL21 did not produce H₂even when formate was added as substrate for formate hydrogenlyase (FHL) pathway. The majority of recombinant protein was produced as an insoluble form, with translocation of a small fraction to the membrane. However, the membrane fraction displayed high activity (~65% of total cell fraction), based on unit protein mass. Supplement of nickel and iron to media showed these metals contribute essentially to the function of [NiFe]-hydrogenase 1 as components of catalytic site. In addition, purified <it>E. coli </it>[NiFe]-hydrogenase 1 using his₆-tag displayed oxygen-tolerant activity of ~12 nmol H₂/(min·mg protein) under a normal aeration environment, compared to [FeFe]-hydrogenase, which remains inactive under this condition.</p> <p>Conclusions</p> <p>This is the first report on physiological function of <it>E. coli </it>[NiFe]-hydrogenase 1 for H₂production. We found that [NiFe]-hydrogenase 1 has H₂production ability even under the existence of oxygen. This oxygen-tolerant property is a significant advantage because it is not necessary to protect the H₂production process from oxygen. Therefore, we propose that [NiFe]-hydrogenase can be successfully applied as an efficient biohydrogen production tool under micro-aerobic conditions.</p

Association Between Knowledge and Attitude About Aging and Life Satisfaction Among Older Koreans

SummaryPurposeThe purpose of this study was to measure knowledge and attitude about aging and life satisfaction among older Korean adults and to examine the influence of attitude and knowledge about aging on these adults' life satisfaction.MethodsA cross-sectional and correlational design was used. A total of 405 older adults were surveyed using a written questionnaire at six elderly welfare centers in a metropolitan city in South Korea in mid April 2011. The data were analyzed using descriptive statistics, Pearson's correlation analysis, and multiple regression.ResultsThe score of knowledge on aging was below the medium level. Attitude about aging was neutral, and life satisfaction was at the medium level. Variables such as female sex, age, economic status, monthly allowance, living with a spouse, self-rated overall health, knowledge and attitude about aging accounted for 33.8% of the total variance in predicting life satisfaction of the older adults. Older age and lower economic status reduced life satisfaction. Being female, having a monthly income of 300,000 Korea Republic Won or more, living with a spouse, and better knowledge and attitude about aging were associated with enhanced life satisfaction.ConclusionThe effect of knowledge and attitude about aging on life satisfaction is significant. Economic status, living with a spouse and self-rated overall health status are also predictive factors in life satisfaction. Consequently, nursing interventions for education and psychological support to increase knowledge about aging and induce a positive attitude towards it should be developed to improve older adults' life satisfaction

Control of nacre biomineralization by Pif80 in pearl oyster

Molluscan nacre is a fascinating biomineral consisting of a highly organized calcium carbonate composite that provides unique fracture toughness and an iridescent color. Organisms elaborately control biomineralization using organic macromolecules. We propose the involvement of the matrix protein Pif80 from the pearl oyster Pinctada fucata in the development of the inorganic phase during nacre biomineralization, based on experiments using the recombinant form of Pif80. Through interactions with calcium ions, Pif80 participates in the formation of polymer-induced liquid precursor-like amorphous calcium carbonate granules and stabilizes these granules by forming calcium ion-induced coacervates. At the calcification site, the disruption of Pif80 coacervates destabilizes the amorphous mineral precursors, resulting in the growth of a crystalline structure. The redissolved Pif80 controls the growth of aragonite on the polysaccharide substrate, which contributes to the formation of polygonal tablet structure of nacre. Our findings provide insight into the use of organic macromolecules by living organisms in biomineralization.117Ysciescopu

Optimization of isoprene production using a metabolically engineered Escherichia Coli

The volatile C5 hydrocarbon, isoprene is an important platform chemical, which has been used in the manufacture of synthetic rubber for tires and also has the potential for various other applications such as elastomers and adhesives. Moreover, isoprene is convertible to biofuel blend stocks such as C10 gasoline, C15 diesel, and jet fuels because of its higher energy content than other biofuels. Although isoprene is currently derived from petroleum, its sustainable supply has been suffered from price fluctuation of crude oil, high refining cost and energy consumption, and low recovery yield of pure isoprene. As an alternative, the biologically produced isoprene (bio-isoprene) has been developed rapidly for the last decade. Bio-isoprene is synthesized from dimethylallyl diphosphate (DMAPP), which is derived from mevalonate (MVA) pathway or the methylerythritol phosphate (MEP) pathway, by isoprene synthase. In this study, metabolic engineering for enhanced production of bio-isoprene was performed by deletion of relevant genes and optimization of culture condition. In comparison of isoprene production between E.coli DH5α and MG1655, lower isoprene production was observed in MG1655. The lower isoprene production in E. coli MG1655 was ascribed to the presence of recA gene which is absent in the DH5α strain. The deletion of recA gene in E.coli MG1655 allows higher isoprene production than E. coli DH5α. Moreover, the optimized expression of isoprene synthesis pathway with 0.03mM IPTG induction enhanced the isoprene production up to 2,850 mg/L. Overall, isoprene production through the optimization was improved by 28.5-fold compared to the initial production of MG1655 strain. Please click Additional Files below to see the full abstract

Engineering de novo disulfide bond in bacterial alpha-type carbonic anhydrase for thermostable carbon sequestration

Exploiting carbonic anhydrase (CA), an enzyme that rapidly catalyzes carbon dioxide hydration, is an attractive biomimetic route for carbon sequestration due to its environmental compatibility and potential economic viability. However, the industrial applications of CA are strongly hampered by the unstable nature of enzymes. In this work, we introduced in silico designed, de novo disulfide bond in a bacterial alpha-type CA to enhance thermostability. Three variants were selected and expressed in Escherichia coli with an additional disulfide bridge. One of the variants showed great enhancement in terms of both kinetic and thermodynamic stabilities. This improvement could be attributed to the loss of conformational entropy of the unfolded state, showing increased rigidity. The variant showed an upward-shifted optimal temperature and appeared to be thermoactivated, which compensated for the lowered activity at 25 degrees C. Collectively, the variant constructed by the rapid and effective de novo disulfide engineering can be used as an efficient biocatalyst for carbon sequestration under high temperature conditions.1194Ysciescopu

Selectively tunable optical Stark effect of anisotropic excitons in atomically thin ReS2

The optical Stark effect is a coherent light-matter interaction describing the modification of quantum states by non-resonant light illumination in atoms, solids and nanostructures. Researchers have strived to utilize this effect to control exciton states, aiming to realize ultra-high-speed optical switches and modulators. However, most studies have focused on the optical Stark effect of only the lowest exciton state due to lack of energy selectivity, resulting in low degree-of-freedom devices. Here, by applying a linearly polarized laser pulse to few-layer ReS2, where reduced symmetry leads to strong in-plane anisotropy of excitons, we control the optical Stark shift of two energetically separated exciton states. Especially, we selectively tune the Stark effect of an individual state with varying light polarization. This is possible because each state has a completely distinct dependence on light polarization due to different excitonic transition dipole moments. Our finding provides a methodology for energy-selective control of exciton states.111612Ysciescopu