Iterative mutagenesis induced by atmospheric and room temperature plasma treatment under multiple selection pressures for the improvement of coenzyme Q10 production by Rhodobacter sphaeroides

CoQ10, which has been widely applied in medicine by dietary supplement, possesses important functions in antioxidant process and bioenergy generation. Iterative mutagenesis introduced by atmospheric and room temperature plasma (ARTP) treatment was studied to improve the coenzyme Q10 (CoQ10) production of Rhodobacter sphaeroides (R. sphaeroides), and multiple selection pressures including vitamin K3 (VK3), Na2S and benzoic acid (BA) were adopted for the first time. After two rounds of mutation and screening, a mutant strain R.S 17 was obtained, and the product titer was increased by 80.37%. The CoQ10 titer and cell density reached 236.7 mg L−1 and 57.09 g L−1, respectively, in the fed-batch fermentation, and the CoQ10 content was 22.1% higher than that of the parent strain. In addition, the spectral scanning results indicated the metabolic flux improvement contributing to the CoQ10 production in R.S 17, and the genetic stability was validated. Based on the iterative mutagenesis introduced by ARTP under multiple selection pressures, the promotion of CoQ10 production by R. sphaeroides was achieved. The significant improvement in fermentation performances and the good genetic stability of R.S 17 indicate a potential way for the efficient biosynthesis of CoQ10.

Mechanism of PQQ and CoQ10 overproduction in Methylobacterium as revealed by comparative genomic analysis between mutant and wild-type strains

Background The microbial synthesis of pyrroloquinoline quinone (PQQ) and Coenzyme Q10 (CoQ10) remains the most promising industrial production route. Methylobacterium has been used to generate PQQ and other value-added chemicals from cheap carbon feedstocks.However, the low PQQ and CoQ10 production capacity of the Methylobacterium strains is a major limitation The regulation mechanism for PQQ and CoQ10 biosynthesis in this strain has also not been fully elucidated. Results Methylobacterium sp. CLZ strain was isolated from soil contaminated with chemical wastewater, which can simultaneously produce PQQ, CoQ10, and carotenoids by using cheap methanol as carbon source. We investigated a mutant strain NI91, which increased the PQQ and CoQ10 yield by 72.44% and 59.80%, respectively. Whole-genome sequencing of NI91 and wild-type strain CLZ revealed that both contain a 5.28 Mb chromosome. The comparative genomic analysis and validation study revealed that a significant increase in biomass and PQQ production was associated with the base mutations in the methanol dehydrogenase (MDH) synthesis genes, mxaD and mxaJ. The significant increase in CoQ10 production may be associated with the base mutations in dxs gene, a key gene in the MEP/DOXP pathway. Conclusions A PQQ producing strain that simultaneously produces CoQ10 and carotenoids was selected and after ANI analysis, named as Methylobacterium sp. CLZ. After random mutagenesis of this strain, we obtained NI91 strain, which showed increased production of PQQ and CoQ10. Based on comparative genomic analysis of the whole genome of mutant strain NI91 and wild-type strain CLZ, a total of 270 SNPs and InDels events were detected, which provided a reference for subsequent research. The mutations in mxaD, mxaJ and dxs genes may be related to the high yield of PQQ and CoQ10. These findings will enhance our understanding of the PQQ and CoQ10 over-production mechanism in Methylobacterium sp. NI91 at the genomic level. It will also provide useful clues for strain engineering in order to improve the PQQ and CoQ10 production.

Improvement of Coenzyme Q10 Production: Mutagenesis Induced by High Hydrostatic Pressure Treatment and Optimization of Fermentation Conditions

Coenzyme Q10 (CoQ10, ubiquinone), a potent antioxidative dietary supplement, was produced by submerged fermentation usingAgrobacterium tumefaciensinstead of chemical synthesis or solvent extraction.Agrobacterium tumefaciens1.2554 was subjected to mutagenesis using a series of treatments including high hydrostatic pressure (HHP) treatment, UV irradiation, and diethyl sulfate (DES) treatment to obtain mutant strains showing higher CoQ10 production than wild-type strains. A mutant strain PK38 with four genetic markers was isolated: the specific CoQ10 content of the mutant strain increased by 52.83% compared with the original strain. Effects of carbon and nitrogen sources on CoQ10 production with PK38 were studied. Sucrose at concentration of 30 g/l was tested as the best carbon source, and yeast extract at concentration of 30 g/l supplemented with 10 g/l of ammonium sulfate was identified to be the most favorable for CoQ10 production using PK38. Fed-batch culture strategy was then used for increasing production of CoQ10 in 5-l fermentor. Using the exponential feeding fed-batch culture of sucrose, cell growth and CoQ10 formation were significantly improved. With this strategy, the final cell biomass, CoQ10 production, and specific CoQ10 production increased by 126.11, 173.12, and 22.76%, respectively, compared to those of batch culture.

Increasing CoQ10 Production by Rhodopseudomonas palustris J001 Using a Two-Stage Fermentation Process

CoQ10 is used not only as a medicine but also as a food supplement due to its various physiological activities. The production of CoQ10 by microbes is a successful approach for generating large amounts of this natural product. The effects of dissolved oxygen (DO) contents and the two-stage fermentation process on cell growth and CoQ10 production by Rhodopseudomonas palustris J001 were investigated. The optimal DO contents for cell growth and CoQ10 production were 45% and 15%, respectively. A two-stage fermentation process, which consists of a 1st stage with 45% DO, a 2nd stage with 15% DO and a synchronous feeding of 2.0% NaAc at the switching time (42 h after inoculation), has proven to be the optimum fermentation process for the production of CoQ10. The maximum biomass, CoQ10 production and CoQ10 production rate were 1.31 g l-1, 89.1 mg l-1, and 1.142 mg l-1 h-1, respectively, increased by 28%, 585% and 426% as compared to the one-stage batch production with 45% DO. The DO level was the major factor to increase the CoQ10 production by the two-stage process.