Project description:Ectoine has gained considerable attention as a high-value chemical with significant application potential and market demand. This study aimed to increase ectoine yields by blocking the metabolic shunt pathway of L-aspartate-4-semialdehyde, the precursor substrate in ectoine synthesis. The homoserine dehydrogenase encoded by hom in H. campaniensis strain XH26 is responsible for the metabolic shunt of L-aspartate-4-semialdehyde to glycine. CRISPR/Cas9 technology was used to seamlessly knockout hom, blocking the metabolic shunt pathway to increase ectoine yields. The ectoine yield of XH26/Δhom was 351.13 mg (g CDW)-1 after 48 h of incubation in 500 mL shake flasks using optimal medium with 1.5 mol L-1 NaCl, which was significantly higher than the 239.18 mg (g CDW)-1 of the wild-type strain. Additionally, the absence of the ectoine metabolic shunt pathway affects betaine synthesis, and thus the betaine yields of XH26/Δhom was 19.98 mg (g CDW)-1, considerably lower than the 69.58 mg (g CDW)-1 of the wild-type strain. Batch fermentation parameters were optimized, and the wild-type strain and XH26/Δhom were fermented in 3 L fermenters, resulting in a high ectoine yield of 587.09 mg (g CDW)-1 for the defective strain, which was significantly greater than the ectoine yield of 385.03 mg (g CDW)-1 of the wild-type strain. This study showed that blocking the metabolic shunt of synthetic substrates effectively increases ectoine production, and a reduction in the competitively compatible solute betaine appears to promote increased ectoine synthesis.
Project description:Biomanufacturing remains financially uncompetitive with the lower cost but higher carbon emitting hydrocarbon based chemical industry. Novel chassis organisms may enable cost reductions with respect to traditional chassis such as E. coli and so open an economic rout to low emission biomanufacturing. Extremophile bacteria exemplify that potential. Salt tolerant halomonas species thrive in conditions inimical to other organisms. Their adoption would eliminate the cost of sterilising equipment. Novel chassis are inevitably poorly understood in comparison to established organisms. Rapid characterisation and community data sharing will facilitate organisms’ adoption for biomanufacturing. This paper describes baseline proteomics data set for Halomonas bluephagenesis TD01 under active development for biomanufactoring. The data record comprises a newly sequenced genome for the organism; evidence for expression of 1150 proteins (30% of the proteome) including baseline quantification of 1050 proteins (27% of the proteome) and a spectral library enabling re-use for targeted proteomics assays. Protein data is annotated with KEGG Orthology enabling rapid matching of quantitative data to pathways of interest to biomanufacturing.