Optimization of inorganic carbon sources to improve the carbon fixation efficiency of the non-photosynthetic microbial community with different electron donors.

As the non-photosynthetic microbial community (NPMC) isolated from seawaters utilized inorganic carbon sources for carbon fixation, the concentrations and ratios of Na2CO3, NaHCO3, and CO2 were optimized by response surface methodology design. With H2 as the electron donor, the optimal carbon sources were 270 mg/L Na2CO3, 580 mg/L NaHCO3, and 120 mg/L CO2. The carbon fixation efficiency in response to total organic carbon (TOC) was up to 30.59 mg/L with optimal carbon sources, which was about 50% higher than that obtained with CO2 as the sole carbon source. The mixture of inorganic carbon sources developed a buffer system to prevent acidification or alkalization of the medium caused by CO2 or Na2CO3, respectively. Furthermore, CO2 and HCO3(-), the starting points of carbon fixation in the pathways of Calvin-Benson-Bassham and 3-hydroxypropionate cycles, were provided by the carbon source structure to facilitate carbon fixation by NPMC. However, in the presence of mixed electron donors composed of 1.25% Na2S, 0.50% Na2S2O3, and 0.457% NaNO2, the carbon source structure did not exhibit significant improvement in the carbon fixation efficiency, when compared with that achieved with CO2 as the sole carbon source. The positive effect of mixed electron donors on inorganic carbon fixation was much higher than that of the carbon source structure. Nevertheless, the carbon source structure could be used as an alternative to CO2 when using NPMC to fix carbon in industrial processes.

Mixed electron donors synergistically enhance CO2 fixation of non-photosynthetic microorganism communities through optimizing community structure to promote cbb gene transcription.

Studies have shown that mixed electron donors (MEDs) can enhance the CO2-fixing efficiency of non-photosynthetic microbial communities (NPMCs), even up to the level of fixation observed when H2 is used as an electron donor. However, this promotion effect is not stable because its mechanism remains unclear. To elucidate the mechanisms involved, allowing further regulation and optimization of the MED system for improving the CO2-fixing efficiency of NPMCs consistently, cbb gene transcription level and efficiency, extracellular free organic carbon (EFOC) content as well as microbial structure of NPMCs under MED and other electron donor systems were investigated. MEDs synergistically promoted CO2 fixation efficiency of NPMCs, even producing levels seen when H2 was used as the electron donor. Subsequent experiments revealed that the cbb gene abundance and transcription level in the MED system were high compared with those in other single-electron donor systems; the concentration of EFOC per unit cell was relatively lower than that in any other electron donor system; and the system developed a large number of dominant heterotrophic bacteria such as Enterobacteriaceae and Vibrionaceae. Data analysis revealed a high negative correlation between EFOC concentration per unit cell and cbb gene abundance as well as gene transcription level. These results implied that MEDs can promote a complex microbial community structure enriched with high-efficiency heterotrophic bacteria, which can effectively reduce excessive EFOC generated by NPMCs in the CO2 fixation process, promoting overall cbb gene abundance and transcription level within the NPMC and thus enhancing CO2 fixation.

An intrabody specific for the nucleophosmin carboxy-terminal mutant and fused to a nuclear localization sequence binds its antigen but fails to relocate it in the nucleus.

The cytoplasmic accumulation of NPM1 (NPMc+) is found in acute myeloid leukemia (AML) with NPM1 mutation. NPM1 must shuttle between nucleus and cytoplasm to assure physiological protein synthesis and, therefore, the elimination of NPMc+ is not a suitable therapeutic option. We isolated, characterized, and produced a functional scFv intrabody fused to nuclear localization signal(s) (NLS) that does not recognize NPM1 but binds to the mutant-specific C-terminal NES (nuclear export signal) of NPMc+, responsible for its cytoplasmic accumulation. The scFv-NLS fusion accumulated in the nuclei of wild type cells and strongly bound to its antigen in the cytoplasm of NPMc+ expressing cells. However, it failed to relocate the majority of NPMc+ in the nucleus, even when fused to four NLS. Our results show the technical feasibility of producing recombinant intrabodies with defined sub-cellular targeting and nuclear accumulation but the lack of information concerning the features that confer variable strength to the signal peptides impairs the development of biomolecules able to counteract pathological sub-cellular distribution of shuttling proteins.