Novel Redox Potential-Based Screening Strategy for Rapid Isolation of Klebsiella pneumoniae Mutants with Enhanced 1,3-Propanediol-Producing Capability

Abstract

This report describes a novel redox potential (oxidoreduction potential [ORP])-based screening strategy for the isolation of mutants of Klebsiella pneumoniae which have an increased ability to produce 1,3-propanediol (1,3-PD). This method can be described as follows: first, to determine an ORP range which is preferred for the wild-type strain to grow and to produce 1,3-PD; second, to subject a chemically mutagenized culture to a reduced ORP level which is deleterious for the wild-type strain. Colonies that showed high specific growth rates at deleterious ORP levels were selected, and their abilities to produce 1,3-PD were investigated. In an ORP-based screening experiment where the ORP was controlled at ؊280 mV, 4 out of 11 isolated strains were recognized as positive mutant strains. A mutant which is capable of producing higher concentrations of 1,3-PD was subjected to fed-batch fermentations for further characterization. Its preferred ORP level (؊280 mV) was significantly lower than that of its parent (؊190 mV). The highest 1,3-PD concentration of the mutant was 915 mmol liter ؊1 , which was 63.1% higher than that of the parent. Metabolic-flux analysis suggested that the intracellular reductive branch of the mutant was strengthened, which improved 1,3-PD biosynthesis. The procedure and results presented here provide a novel method of screening for strains with improved product formation.

Assessing the potential of mutational strategies to elicit new phenotypes in industrial strains

“…A major remaining challenge is engineering environmental tolerance phenotypes to allow robust growth and production under adverse industrial conditions (6, 7). In general, these phenotypes are hard to engineer, because stress conditions elicit multigenic responses (8) coordinated at the transcriptomic and proteomic levels (9, 10).Robust industrial strains have been traditionally obtained through adaptation and selection (11,12). Because natural adaptation is time-and resource-intensive, mutagens are used to introduce diversity in the population and accelerate the process (13).…”

“…Robust industrial strains have been traditionally obtained through adaptation and selection (11,12). Because natural adaptation is time-and resource-intensive, mutagens are used to introduce diversity in the population and accelerate the process (13).…”

Genetically Engineered Strains: Application and Advances for 1,3-Propanediol Production from Glycerol

“…Technologies like atmospheric and room temperature plasma have been used to produce new strains, and good results were obtained (52). Therefore, complex agents or other violent factors are a promising choice.…”

“…Compared with parent strain, the production of the two mutants LM03 and LM05 was improved by 33 and 30 % respectively. A large number of studies suggests that the rate of positive mutation by LiCl is much higher than the single UV mutagenesis, and protoplast mutagenesis by UV combined with LiCl is more liable than mycelium mutagenesis to obtain the high yield strains (52,58,65,66). N-methyl-N'-nitro-N-nitrosoguanidine N-methyl-N'-nitro-N-nitrosoguanidine (NTG) is an alkylating mutagenesis agent, which is a commonly used chemical mutagen.…”

“…LiCl is usually used in combination with UV mutagenesis in breeding. Du et al (52) obtained mutants by UV light and LiCl mutagenesis derived from the parent strain of K. pneumoniae. In this study, a novel redox potential (oxidoreduction potential [ORP])-based screening strategy was designed for the isolation of mutants with high 1,3-PD production.…”